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KB-0002-Jun08 ISPE Fundamental Knowledge Brief Biotechnology Basics The information contained in this Knowledge Brief was adapted from training materials as part of the ISPE Training Course on Biotechnology Basics, taught by Jeff Odum. Introduction Increasingly, the pharmaceutical industry has undergone a shift in emphasis from traditional chemical based medicines to those involving large molecules. The large molecule medicinal is a direct result of the major role of biotechnology in our industry. “Biotechnology is a combination of advances in our understanding of molecular and cell biology and human genetics, and how the human immune system fights disease.” This modern definition is changing rapidly as the technologies continue to develop and advance within the industry. For obvious and good reason, a major application of biotechnology is toward human health care, including: the detection and treatment of diseases; human growth; and vaccines. Biotechnology holds tremendous potential in other applications that have www.ISPE.org © Copyright ISPE 2008. All rights reserved. major economic and social impact in our society, such as agriculture, food processing, fuels, and wastewater management. The focus of this Knowledge Brief will be on biotechnology’s application toward human therapeutics. The general information provided here is drawn from ISPE training courses that are focused on biotechnology and biomanufacturing processes. The purpose of this Brief is to provide basic concepts explaining the science of biotechnology and how science and process are combined to lead to the manufacture of a human therapeutic product. More detailed information can be found in ISPE publications and in select ISPE training programs available on-site or on-line. The Science of Biotechnology Cells are the fundamental working units of all living things. Within the cell are all the instructions needed to direct the cells activities. These instructions are stored in deoxyribonucleic acid (DNA) in the nucleus of cells. ISPE Knowledge Brief Page 2 Biotechnology Basics only tiny quantities can be extracted from human tissue. A significant and worthwhile goal of biotechnology is the production of sufficient amounts of high quality human proteins. The structure of the DNA molecule is identical among all living things, from an amoeba to a killer whale, from a blade of grass to a redwood tree. This is what makes recombinant DNA technology – the transplanting and combining of genetic information, one organism to another – possible. The term recombinant DNA technology is more popularly known as genetic engineering. Bioprocess Basics Now let’s take a look at how the science and the process integrate to lead to the manufacture of a human therapeutic product. Figure 1. Human Genome Program, U.S. Department of Energy, Genomics and It’s Impact on Medicine and Society: A 2001 Primer, 2001. DNA is a long molecule that is composed of a sugar, phosphate, and one of the following four nitrogen bases: • • • • Adenine (A) Thymine (T) Guanine (G) Cytosine (C) A sugar, phosphate, and base together constitute a nucleotide. These nucleotides pair up into strands that twist together to form a double helix. A segment of DNA, in conjunction with A process is defined as any operation or series of operations by which a particular objective is accomplished. In this case, the objective is the production of a biotech drug product. some proteins, is a chromosome. Each chromosome contains many genes. Genes contain instructions for making proteins. Proteins are the major structural and regulatory molecules essential for life. Proteins act alone or in complexes to perform a particular function in the body. Human proteins are valuable in treating disease. For example, diseases caused by a protein deficiency can be treated with the human protein itself. However, The DNA Code In Figure 2, the “basic” biotech process steps are shown as upstream and downstream operations. Upstream operations can be thought of as cell growth and separation. Downstream operations are product purification and filling. Almost every biotech process can be sub-divided into these two basic elements. The basic unit operations are shown in Figure 3. These unit operations represent the “standard” operations that most companies implement in their manufacturing operations. The basic biotech process elements are: The DNA alphabet is A, G, C, and T. The sequence of these four letters determines DNA information. A DNA sequence is actually a string of three-letter “words.” AGCTTCCGATCGGTA actually reads: AGC TTC CGA TCG GTA. Each three letter sequence, or condon, specifies one of 20 amino acids. Amino acids are the subunits of protein. AGC is the code for the amino acid Serine and TTC is the code for Phenylalanine. No matter where they are found, the condons are always the same for the same amino acid. • • • • • Bioanalysis Fermentation Separation Purification Filling Bioanalysis Nearly every process conducted in a biopharmaceutical company requires © Copyright ISPE 2008. All rights reserved. ISPE Knowledge Brief Page 3 Biotechnology Basics number of different options that companies have as a method of breaking down cellular material: • Nonmechanical - Freezing - Detergents - Enzymes • High pressure - Centrifugation • Homogenization • Mechanical grinding Most common in use today is centrifugation. Centrifuges are one of the most common equipment items used for cell disruption. A GMP model can cost well over $500,000 (US). The downstream operations begin by separating the “good” from the “waste” in the product materials. Separating is nothing more than a filtration operation to accomplish this activity. Figure 2. “back up.” The analysis phase of manufacturing is critical as proof of the drug’s safety, purity, and efficacy. Analytical Methods Back up regulatory submissions. • Support pre-clinical and clinical studies. • Monitor environmental conditions during manufacturing. • Monitor quality of the manufacturing process. Cell culture is a specific type of fermentation that involves the process of taking cells from living organisms and growing them under controlled conditions. Cell culture is part of the upstream processing operations; literally engineering and growing the cell line to be used to manufacture the drug product. The separation steps are: Once fermentation is complete, the desired product must be recovered, separated out, and purified. Purification Fermentation Separation Fermentation is the process by which living cells obtain energy through the breakdown of glucose and other molecules. Fermentation refers to the large-scale cultivation of microorganisms. Recovery is the separation of crude product from microbial mass and other solids and liquid medium, to prepare it for purification. Figure 3. Basic Processing Steps. © Copyright ISPE 2008. All rights reserved. Product recovery usually requires some type of cell disruption. There are a • Extraction and precipitation • Filtration - Microfiltration - Ultrafiltration Filtration skids are highly automated and can cost in excess of $250,000 (US). The purification steps are: • Chromatography - Gel filtration - Ion exchange - Hydrophobic interaction (HIC) - Affinity The purification steps are high risk and very costly to perform. Formulation of the drug is a critical step that is very important to the patient. ISPE Knowledge Brief Page 4 Biotechnology Basics The required protein must be modified to a stable, sterile form that can be taken by the patient. Remember, proteins cannot be taken as pills because they are broken down in the stomach. Until recently, biotech products were exclusively sterile injectibles. Now, inhalation and transdermal delivery options give patients greater flexibility. Filling Filling is the process of putting the drug product into a container. Two general categories of filling are: • Bulk • Final Bulk filling is defined as the placement of larger quantities (5L-100L) of product into containers for shipment/storage. Some examples of containers are: • • • • Vials Ampoules Syringes Dental cartridges Final filling is defined as the placement of drug product into its final container/ closure system. The majority of production facilities produce product in bulk. Many companies ship their bulk to contract filling firms. Conclusion This document is a basic review of the general principles of biotechnology and biotech processing. As such it is intended to serve as a primer on this highly sophisticated area of science and technology. For more detailed information, you can search ISPE articles and publications on this subject, including the ISPE Baseline® Pharmaceutical Engineering Guide Series: Volume 6: Biopharmaceutical Manufacturing Facilities. For the most current information and up-to-the-minute discussions on biotechnology and biotech processing, visit our Biotechnology Community of Practice on the ISPE Web site. About the Author Jeffery Odum is president of NCBioSource USA and ISPE’s North American Continuing Education Advisor. He has been involved in the biopharmaceutical industry for more than 20 years with expertise in engineering design, construction, and plant operations. Prior to founding NCBioSource, he was a successful market leader for an international engineering design firm specializing in biopharmaceutical manufacturing facility design and construction. His experience includes design and construction of many of the industry’s major manufacturing projects, as well as consulting roles for a number of the global biotechnology industry leaders. Odum is a nationally recognized author and speaker with industry insight in the areas of regulatory compliance, facilities and process design, and project management for biopharmaceutical companies. He has been a Member of ISPE for more than 15 years and has been a presenter and course leader for more than 25 professional training and education sessions within the industry. Odum is a past chairman of ISPE’s North American Continuing Education Committee (NAEC) and the ISPE Training Committee. As North American Continuing Education Advisor, he works closely with the NAEC and the Director of Continuing Education on development of the society’s North American conferences. He is also a Member of ISPE’s Technical Training Staff. In 2002, Odum received the Society’s prestigious Richard B. Purdy Award for Outstanding Achievement within the biopharmaceutical industry. Odum is the author of more than 25 published works on many critical issues, including process improvement and execution to meet regulatory guidelines issued by the FDA and other international regulatory bodies. He was also one of the lead chapter authors for ISPE’s Biopharmaceutical Manufacturing Facilities Baseline® Guide. Odum graduated from Tennessee Technological University with a degree in mechanical engineering and received his Masters Degree in engineering from the University of Tennessee-Knoxville. • © Copyright ISPE 2008. All rights reserved.