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We live in a climate of fear.

For more than two years, the world has held its breath, waiting for the proverbial other shoe to fall. American soldiers have been deployed to Afghanistan and Iraq, searching for terrorists, threats to global security and the leaders of tyrannical regimes. It is not what these soldiers have found that has instilled this sense of fear in the national consciousness, but rather, what they have not

Included stories:

Raising the Bar

Engineering for the Body
found—weapons of mass destruction, biological agents, crude yet effective tools of terror we know exist, whether or not we see them. At the University of Nebraska-–Lincoln there is the sense that we are far removed from these spoken yet unseen dangers. This is the heartland, safely ensconced within the center of the most powerful country in the world. And yet, it is here that a crucial front in the fight against terrorism is taking place, within the laboratories of the Biological Process Development Facility (BPDF), led by director Michael Meagher, Donald F. Othmer professor of chemical engineering. “I am pleased we are in a position to contribute to the protection of our nation in the defense against bioterrorism,” said Prem Paul, UNL vice chancellor for research.

The history of biological warfare has been documented as early as 1346 when during a siege on the port of Caffa on the Crimean peninsula, soldiers threw bodies infected with plague over the city walls. Attempts at biological warfare quickly became more sophisticated. More than 60 years ago, the United States began to develop and use the botulinum toxin as a biological weapon. Although the American bioweapons program ended under Nixon’s orders in 1970, biowarfare research continued in other parts of the world. After the 1991 Gulf War, Iraq admitted to the United Nations to having produced 19,000 liters of botulinum toxin—toxin that still is unaccounted for and is enough to kill the world’s population three times over. As of yet, there is no vaccine available. The threat of biological terrorism casts a wide shadow, and the American people are left wondering whether the military is capable of effectively dealing with the consequences of a biological terror attack on American soil. “The best scenario is that we never have to use these vaccines,” Meagher said. “But the American people will not tolerate our government not being prepared.”

Today, botulism, caused by the botulinum neurotoxin, is one of the most feared biological agents in the world. The toxin has seven serotypes each of which require a separate vaccine. Forms of botulism are classified by the mode of exposure and include inhalational botulism, food-borne botulism and wound botulism. Symptoms include blurred vision, dysphasia and dry mouth and muscle weakness, which leads to paralysis and eventually, muscle failure. The severity depends upon exposure and any delay before antitoxin therapy. It is perhaps ironic then, that the botulinum toxin is also the first biological toxin that has been licensed for medical treatment, in forms such as the commercial Botox, used to treat frown lines. The processes Meagher and his research team are developing will enable the production of vaccines to protect against botulism serotypes A, B, C, E and F within one to two years, and serotypes D and G within five years. “We’re going to make life difficult for those trying to come up with new strains of biological agents,” Meagher said.

Fermentation processes have been used since 4,000 B.C. when the Chinese discovered a range of processes including the use of lactic acid bacteria to make yogurt. Thousands of years later, Louis Pasteur came to the realization that if germs were the cause of fermentation, they could also be the cause of contagious diseases. He began work on a germ theory that eventually led to his work on vaccinations. In a far more sophisticated manner, Meagher’s research follows the same trajectory. More specifically, Meagher’s research focuses on developing biological processes specific to producing therapeutic recombinant proteins—a molecule that will assist the body in improving its health. To do this, he uses Pichia pastoris, a methyltrophic yeast that is flexible and can be used for protein production. Pichia pastoris is ideal in that it offers high expression levels, can be easily adapted to large-scale fermentation for the production of recombinant proteins and is less expensive than expression in insect or mammalian systems. The challenge lies in finding the best conditions for producing each protein and assuring a consistent, reproducible and reliable process. Once a process has been successfully developed, researchers provide clients with detailed documentation so they can use the fermentation process for their own needs. “We can develop any recombinant molecule a client wants. We have just about all the expression systems now—transgenic plants and animals, mammalian cells, bacteria and yeast—we can do it all,” Meagher said.

The fermentation processes, however, are only a small part of the puzzle. All of the elements of the BPDF, are equally important in developing a vaccine or any recombinant product. It all starts in the Molecular Biology Laboratory with development of the best recombinant strain of Pichia pastoris that will express the correct therapeutic protein, or vaccine, at very high levels. Then the new strain is transferred to the Fermentation Development Laboratory to develop a fermentation process that provides starting material for the Purification Development Laboratory which has the responsibility for developing the purification process itself.

Scott Johnson, manager of the Purification Pilot Plant, prepares a chromatography column. Each column is packed with resin to purify a protein.
The end result from the purification development activities is a process that is simple, scalable and reliable enough to make a product consistently at the 99.5% purity level. The first 10 mg of recombinant protein produced by the Purification Development Laboratory is delivered to the Analytical Methods Development Laboratory which is responsible for the analytical methods that are used to quantitate the vaccine during all aspects of process development, from fermentation to the final purification step. Once the analytical methods are developed they are transferred to the Quality Control Laboratory, which is responsible for the routine analysis. After the process and associated analytical methods are developed, the entire package is transferred to the Current Good Manufacturing Practices (cGMP) staff, who will produce the protein in the BPDF fermentation and purification pilot plants. The quality assurance staff also is very critical as they have the responsibility to oversee the generation of the Production Batch Records, a sophisticated cookbook of the entire process, and the entire process from a quality perspective.

The BPDF has been in operation since 1990. The facility works with public and private sector clients on process research and the early manufacturing of new therapeutic molecules for human clinical testing. Seven work groups, divided among development, production and quality, work to provide complete process development, technology transfer and the manufacture of Phase I clinical material under cGMP. “If you’re going to be good you have to be unique—do something that no one else has done,” Meagher said. Though currently housed in Filley Hall on East Campus, the BPDF will be moving to Othmer Hall in December 2003. The third floor will house research and development facilities including research labs for Meagher, William Velander, chair of the Department of Chemical Engineering, and Anu Subramanian, assistant professor of chemical engineering. The basement, when completed in 2005, will house a pilot plant capable of producing clinical research materials for use in human trials. “Speed to market is critical,” Meagher said. “Once the construction is complete we will be able to take a process from beginning to end within our own facilities.”

Dr. Shinichi Taoka, Analytical Methods Laboratory, injects a sample into the injection port of a high performance liquid chromatograph (HPLC), which is used in analyzing samples from the botulism research.
The BPDF is entirely self-sufficient and recently benefited from an $11 million grant from the National Institute of Allergy and Infectious Diseases, a division of the National Institutes of Health. The University will receive $6.5 million. The grant program emphasizes partnerships between universities, government and the private sector to speed up the development of products needed for biodefense. To that end, researchers at UNL will be collaborating with the other recipients of the grant: DynPort Vaccine Co., the U.S. Army Medical Research Institute of Infectious Diseases and HTD Biosystems Inc. The funds will enable Meagher and his team to continue their research and development of fermentation and purification processes to enable the creation of a heptavalent botulism vaccine.

“One of the many benefits that will emerge from this research is that we’re going to learn a lot about infectious diseases, which will be tremendous in terms of parallel applications of military technology,” Meagher said. But more than that, the grant speaks to the strength of the BPDF, which is the only facility of its kind at any American university. “Before you can say who you are, you have to build what you say you are,” Meagher said. “And we strive to be recognized as the best.”

In addition to federal grant projects, Meagher and his team work with private industry clients. “Other projects we’ve worked on include developing processes for therapeutic proteins to combat multiple sclerosis, cancer, hepatitis and cystic fibrosis,” Meagher said. It takes anywhere from five to eight years for a product to become commercially viable at a cost of about $800 million per drug. Only one in 10 will succeed. “When you do process development, you need anywhere from 100 milligrams to one gram of protein. It’s a very expensive endeavor,” he said. Meagher’s work doesn’t guarantee success, but while he and his team hope that the eventual product succeeds, they do not have to be as invested in the end result. The work of the BPDF strictly involves helping clients develop processes to make new drugs. The responsibilities of initial discovery and using the developed process to manufacture a successful product remain with the client.

Dr. Todd Swanson, research assistant professor and manager of the Analytical Methods Laboratory and the Quality Control Laboratory, operates the HPLC from a remote computer.
The BPDF is having a significant impact in other areas. “These facilities provide a place where we can do solid research in process development, educate students and at the same time serve as an economic engine for the state,” Meagher said. “And that to me embodies what a university is supposed to do.” The BPDF provides an excellent hands-on opportunity to educate Nebraska students in biological processes, thereby strengthening both the graduate and undergraduate programs in chemical engineering. It also enhances the growing biotechnology industry within the state. “Our undergraduate students get significant, industrially relevant experience because our operation is very reflective of conditions in industry. We combine basic science with applied science with applied engineering and everything in between,” Meagher said. “I strongly believe in undergraduates participating in research and I look for highly motivated students. We start them out washing dishes and they work their way up.”

Meagher’s entire staff, in fact, is highly motivated and includes Ardis Barthuli, coordinator of quality assurance, Scott Johnson in the purification pilot plant, and Mehmet Inan in the molecular biology lab. “I have a fantastic staff,” Meagher said. To ensure that the precise work the BPDF staff undertakes is done correctly, Meagher strives to hire the most qualified people in the field. Once employed, each employee, research assistant or student undergoes a comprehensive and formal training program. The quality component of the research is significant, time-consuming yet absolutely necessary to ensure that products created from BPDF processes will do no harm.

Sarah Fanders, research technologist for Dr. Mehmet Inan, research assistant professor and manager of the Molecular Biology Laboratory, works on a project in the lab.
Despite the many successes of the BPDF, Meagher is not slowing down. He has plans for both the present and the future. “There’s a lot going on, including a $5.5 million building project, a $6.5 million NIH contract, 30 people to feed, the basement project on the horizon, as well as serving as the chair of the College biomedical engineering task force. I’m pulled in a lot of directions,” Meagher said. He also has plans for the evolution of the BPDF. “My long term goal is to get 15 to 25 companies to each contribute to an endowment for the facility. Depending on their financial commitment, each of those companies would get a certain level of say on what type of research is going on in the facility. I would also like to see the BPDF become a Center of Excellence,” Meagher said. And yet, helping people remains his priority. “It is tremendous to think about the difference we’re making at Nebraska, and at the University, to fight bioterrorism,” Meagher said.



Raising the Bar

Ardis Barthuli
In an environment where accuracy is non-negotiable, quality assurance assumes an enormous responsibility. In the Biological Process Development Facility (BPDF), Ardis Barthuli, Coordinator of Quality Assurance, is meeting the challenge head on. “Quality is making the product we say we’re going to make, doing it the way we say we’re going to do it, and ensuring that our product won’t be harmful,” Barthuli said.Barthuli and her staff work to ensure that the labs and development facilities of the BPDF meet Good Laboratory Practices (GLP) and current Good Manufacturing Practices (cGMP), systems that laboratories and manufacturers use to build quality into their research and products. Facilities are inspected periodically, both internally and through client audits and FDA inspections. “Audits are a good learning experience because every auditor has a unique perspective,” Barthuli said. Product samples are tested during production by the quality control group to assure that the end product meets client specifications.

To aid in maintaining high standards of quality, Barthuli and her staff built quality into the process. It has taken almost four years to bring the facility into cGMP compliance and at present, 30 percent of the facility’s GMP staff is dedicated to quality assurance. Integrating quality into the facility involves several approaches. The facility recruits highly qualified people with a strong work ethic, establishing a foundation upon which quality can be built. All employees, including graduate and undergraduate students, undergo a comprehensive training program. There are specific standard operating procedures in place on product testing, raw material control, how to operate and clean equipment, and the many other details associated with maintaining compliant facilities. “We need to make sure that everyone does the same thing accurately and consistently,” Barthuli said.

Maintaining quality and adhering to good practices can be difficult. “Our greatest challenge is the constant change within the industry and the technology we’re using. We want to be at the forefront of development so it is imperative to keep up with the change,” Barthuli said. “In a university setting, we have to prioritize what we can and cannot do.” Despite the challenges, Barthuli and her staff look forward to inhabiting the new facilities in Othmer Hall, where many of the limitations they have had to deal with in Filley Hall on East Campus, no longer will exist. The QA staff will have to adapt to the BPDF’s new home and install new procedures for quality assurance, but the entire staff will be in one place, rather than scattered throughout four different floors. The unit will become more centralized and cohesive.

The quality assurance staff also produces the necessary documentation for the successful transfer of products and processes to clients. Barthuli knows her job has been done right when technology transfer is seamless. The work of quality assurance is time-consuming and costly but very necessary. “Quality is extremely important,” Barthuli said. “Our products are being used in human trials and we always keep that in mind.”

Roxane Gay

Engineering for the Body

Suzanne Rohde
Biomedical engineering is, for many, a field of study veiled in mystery. How biomedical research relates to engineering is even further obscured. It is a combination of the physical and the physiological; machines and man. Funding from the National Institutes of Health has increased dramatically over the last ten years as the nation’s population ages and there is an increased demand to study problems from a biomedical perspective. And yet, the newest program of study offered at the graduate level within the College of Engineering & Technology is perhaps the least understood.

In only its second year, the biomedical engineering program is poised to enhance curriculum offerings within the College and is seen as a first step toward a more progressive atmosphere in which departments collaborate for the purpose of interdisciplinary study and research. “The biomedical engineering program joins faculty and students with similar research interests and allows them to draw from a much wider background,” said Suzanne Rohde, professor of mechanical engineering.

Biomedical engineering encompasses many disciplines including engineering, biology and medicine. It endeavors to improve human health through multidisciplinary activities that integrate the engineering sciences with the biomedical sciences and clinical practice. Areas of strength in the UNL program include bio-therapeutics, biomaterials and composites such as drug delivery systems, cellular, tissue and genetic engineering, biosensors and interfaces, biomechanics, systems, computation and information processing, and biomedical instrumentation. “This program is ideal for students whose biomedical interests don’t fit within traditional engineering disciplines,” Rohde said. “And almost every department in the College is lending their expertise to the program.”

A wide range of biomedical engineering-related courses is offered through the College, the University of Nebraska Medical Center and the University of Nebraska at Omaha in subjects ranging from biophysical principles to occupational safety hygiene engineering. The program doesn’t have courses specifically listed as biomedical engineering courses, encouraging students to acquire a specific knowledge base rather than aim for taking a specific set of courses to fulfill degree requirements. Furthermore, Ph.D. candidates pursuing the specialization in biomedical engineering must take 24 credit hours of graduate level engineering courses and at least 12 credit hours of biomedical science courses.

Ultimately, the biomedical program is interested in creating innovative opportunities for engineering students and providing an educational experience that is integrated and interdisciplinary. “We envision scenarios where a student with a biology background is paired with a student in a chemical engineering background to conduct research on biological processes,” Rohde said. “We want to connect students with complementary interests.”

Roxane Gay