DNA: The crystal ball
03/01/2001
Emerging genomic technology could change the way drugs are manufactured and force contamination control to evolve with it
by Mark A. DeSorbo
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Situated in the shadows of Scotland's Edinburgh Castle is The Roslin Institute, the birthplace of the famous cloned sheep, Dolly, and the site where human monoclonal antibodies for melanoma and other cancers are produced inside the eggs of genetically modified chickens.
Across the pond, somewhere in Cambridge, MA, is an old factory where heart-shaped candy boxes were once stamped out. Within this retrofitted building of yesteryear, a futuristic robot named "Zeus" flexes its computer-driven muscles, searching welled genetic material for specific DNA inroads that provide detours to new and more profitable drugs.
Could this be the end of an era, a time when it took scientists many years and millions of dollars to create new medicines and find cures for what ails us?
Most believe genome-based technologies will forever change the way drugs are discovered and administered, along with being a giant leap toward finding cures for diseases that plague the world. Others, who equally embrace discoveries that aim to improve the quality of life, also agree, but recognize that the pharmaceutical and biotechnology industries are in a constant state of flux, an evolution that has agenda items, such as proposed guidance from the U.S. Food and Drug Administration (FDA), that still need to be addressed before new trails are blazed.
Pharmaceutical professionals know this much is true; whether it is in Scotland, the United States or some other distant land, the pharmaceutical industry is undergoing a global transformation, and at the heart of the remedy revolution the core value of contamination control must evolve with it.
Silicon Valley contribution
DNA microarrays are perhaps one way the Silicon Valley is having an impact on the pharmaceutical industry. A typical microarray will have thousands of single-stranded gene fragments that are fastened to a platform, which can be a silicon or glass wafer or a nylon sheet, which are also manufactured in sterile environments.
Microarrays, like those manufactured by Affymetrix (Santa Clara, CA), enable scientists to scan up to 60,000 gene sequences at once. The company produces arrays that aid scientists in deriving drugs from plants as well as from human and animal DNA for toxicology, neurobiology, immunology and other specific research applications using genomes.
Affymetrix also manufactures the actual arrayers, the same machine that one drug maker nicknamed "Zeus." The device is encased in a controlled environment, and within it, a robotic arm reaches into an area about the size of a baby's crib. There, thousands of samples of genetic material sit in tiny wells that have been etched onto plastic plates. The individual samples are identified by a specific bar code, and the arrayer searches these for particular symbols. Once the arrayer finds what it's looking for, it dips a fine needle into a well and extracts a droplet of liquid DNA.
That DNA droplet is then transferred onto a silicon or glass wafer or nylon sheet, and the process starts all over again until the sheet or wafer is filled, forming the microarray. The sheet or wafer is then rolled up, inserted into glass tubes and showered with radioactive dye and genetic material, which ranges from healthy to diseased cells. The exercise yields information that is placed under a fluorescent light, and whatever glows could be a researcher's first step toward a cure for a deadly disease.
"[Arrayers] are bench-top tools, and when they are manufactured they have to be made in a cleanroom," says Dr. Frank Sistare, director of the Center for Drug Evaluation and Research Applied Pharmacology Research Division. "Arrays are also done in a cleanroom. There all kinds of contamination control practices that need to be followed. Temperature, humidity and dust all come into play."
Use of this technology is being carried out at Millennium Pharmaceuticals Inc. (Cambridge, MA) in its efforts to make a better blood pressure drug. When contacted by CleanRooms, officials at Millennium as well as Biogen Inc. (Cambridge, MA) refused to discuss its use of genome-based technology and the contamination control protocols they follow.
Millennium's plan to develop better drugs using the genome, however, was highly publicized in the January 15 issue of Time magazine. According to the article, what would have taken the drug maker 10 years is expected to take just two because of the high-throughput arrayer they call "Zeus."
The yoke's on them
While Millennium carves out its future, genome-technology has plenty of opportunities to go around.
In fact, a report from investment bank UBS Warburg LLC indicates that there are more than 200 companies involved in developing more than 400 monoclonal antibodies. A report from The Boston Globe also indicates that contract manufacturers and drug companies are beginning to commit resources to add manufacturing capacity. Two companies, Immunex Corp. (Seattle) and Biogen already have plants, and the companies are gearing up to start production at the end of next year.
"There will not be any one party that will unlock all the doors," says Mel Rothberg, executive vice president of Viragen Inc. (Plantation, FL). "There are multiple therapeutic possibilities out there."
Viragen, a manufacturer of immunomodulatory therapeutic products, has recently teamed up with The Roslin Institute, a biotechnology center, to develop monoclonal antibodies to fight cancer inside the eggs of genetically modified chickens.
Deemed "The Avian Project," the collaborative scientific effort uses the same technology that was used to clone the famous sheep, Dolly. [See "Dolly the double," CleanRooms, September 1997, p. 1.]
The project, Rothberg explains, has two phases. First, scientists modify the chicken's genetic code to allow for it to produce in its egg white a particular pharmaceutical product. The second step is to clone that bird so that it "cookie cuts, or duplicates itself repeatedly" in order to produce a flock of those birds to produce those eggs.
Because chicken eggs have an inherent defense mechanism against viruses and bacteria, cleanrooms are not needed for the infancy research stages, says Dr. Helen Sang, Roslin's lead scientist for The Avian Project.
"When working with the eggs, we use laminar flow hoods, but research is by no means conducted in a sterile environment," she says. "Chicken eggs are very resistant to bacteria, so you can work with them in fairly crude conditions."
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Dr. Sang says the collaborative effort between The Roslin Institute and Viragen's Scotland facility has not yet produced any compounds for clinical trials. "When we do produce proteins for clinical trials, we'll have to change conditions, and we are hoping for a new facility, which will include cleanrooms, so we can show proteins are produced under sterile conditions," she adds.
As with any type of biopharmaceutical process, absence of bacterial, viral and fungal contamination is essential, for it can inhibit the normal functioning of the cell as well as make modification impossible.
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The environment of future production of Avian Project proteins was compared to that where Dolly's duplication took place. In Dolly's case, her existence came about in ISO Class 7 (Class 10,000) cleanrooms, where specially selected tissue samples were cultured, manipulated and stored. In the other controlled environment, unfertilized donor eggs were recovered, genetically altered for cloning.
Modifying animals to yield treatments for cancer and high blood pressure is not the only benefit of genome-technology. In fact, completing DNA sequences of potentially deadly bacteria allows scientists to dissect pathogenic molecules to determine what makes them tick and kill.
More than two-dozen scientists at the University of Wisconsin-Madison Genome Center believe they are one step closer to learning the secrets and ending the deadly deeds of the E. coli O157:H7 strand. The food-borne pathogen sickens more than 75,000 people and results in numerous deaths annually. It was first identified in 1982 from an outbreak from contaminated hamburger, and reported cases have risen steadily. There are currently no effective treatments for the sickness, which causes a severe form of bloody diarrhea and can also release toxins that damage kidneys and cause renal failure. [See "Second E. coli scare sparks meat recall," CleanRooms, January 2001, p. 4].
Based on the team's findings, the group discovered "islands of pathogenicity" throughout the genome, which the team believes may make it harder to control the public health threat.
In a comparison of the O157:H7 strain and a benign strand, the O157:H7 had 1,300 additional genes that were not found in the harmless strain. The benign "cousin" bacteria also had 530 unique genes that were not shared with O157:H7. What that tells scientists, according to their report, is that the deadly bacteria adapts to its environment and develops resistance to antibiotics.
Those additional genes, scientists believe, can be exchanged across entire families of bacteria, including related organisms like Salmonella and Shigella, the Plague-causing organism Yersinia and the plant pathogen Erwinia.
"We have found that the genomic pieces are constantly shuffling around so that any particular strain contains a subset of the full range available," says Dr. Fred Blattner, director of the university's genome center.
If that full range, or "pathosphere" is large enough, Dr. Blattner says, it could be an underlying factor in the emergence of new diseases.
 The Roslin Institute in Edinburgh, Scotland. |
New genes help explain why E. coli O157:H7 infections are sometimes difficult to treat, says Guy Plunkett III, a university geneticist. Certain antibiotics used to combat E. coli can actually stimulate virally infected bacteria to produce more viruses and viral toxins. "In the course of treating the disease, you could actually exacerbate the problem," Dr. Plunkett says, adding that another set of newly discovered E. coli genes might allow the bacteria to withstand fever, one of the body's defenses against infection.
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Nicole Perna, lead author of the university's report, notes that E. Coli sequencing accomplishment gives scientists real targets for future work on treatments and vaccines. Researchers, she says, need to better understand how these different clusters of genes are being transferred horizontally across different species.
"One of the first things we can do is improve our detection and surveillance before it becomes a public health issue," she says. "We now have a far better distribution of genetic markers to help identify this in the field."
This information, Perna adds, should allow scientists to detect the presence of E. coli more easily, whether it is in humans or potentially contaminated food.
Growing pains
While the promise of an improved quality of life is encouraging, many professionals remain concerned by the apparent lack of harmony that is needed for an industry that is headed in a global direction.
International regulatory issues remain moving targets, says Russell Madsen, senior vice president of science and technology for Parenteral Drug Association (Bethesda, MD).
Whatever the recommended practice or standard may be, Madsen points out there is a U.S. model and a European model, and proposals for new regulations continue to be drafted.
"Australia just came out with new current Good Manufacturing Practices (cGMPs) and Canada has just proposed sterile assurance criteria," he says. "There are all kinds of organizations, and they are all coming up with standards and recommended practices that are pretty much the same, but have differences, and that drives everyone crazy. So people are having a hard time complying. It's just like the Senate and the House; they can't agree."
And the absence of cohesiveness could be regarded as growing pains, for the state of an evolving industry is indicated by what is going on in throughout the world.
For the better part of two years, China has been enforcing a crackdown on its substandard drug system, which has resulted in the shut downs of many pharmaceutical companies. [See "China copes with drug woes," p. 1] And Asia has indeed caught the eye of many drug and medical device manufacturers as a strategic hot spot [See "Schering-Plough fuels Singapore life sciences fire," Feb. 2001, p.1].
China is not the only place in the world that is coping with the need for change. Europe is embroiled in food safety issues that stem from mad cow disease, and along with dealing with numerous outbreaks, the European Union (EU) is busy trying to establish an organization similar to the FDA. [See "Mad cow disease: a shot heard round the world," p. 8.]
The buck stops at the FDA
Tackling standards for DNA-developed drugs would not be prudent at this stage of the game, Dr. Sistare cautions.
"The biggest mistake we could make is making regulations too early," he says. "We need to determine and find out what the best approach is to assure quality, consistency and efficacy. We don't want to put guidance out too early because there are a lot of issues that still need to be resolved. This field is in a rapid state of flux."
The FDA, like any governing agency, has felt the ebb and flow of that unrest. Over the last year, it's been coping with outbreaks of food-borne illnesses. It has dealt with great scrutiny over newly launched programs, like its Hazard Analysis Critical Control Point program. The FDA has also been taking steps to improve the ways and means of drug compounding to improve the safety and efficacy of drugs as well as contain life-threatening, hospital-borne viruses.
On top of that, the agency recently had to answer to a report that indicates that its regulators took nearly 40 percent more time in 2000 than the previous year to approve new prescription drugs, which ended a seven-year streak of faster authorization times and hinted a slowdown.
Last year, the FDA approved 27 medicines that have active ingredients never before sold in the United States. The drugs are classified by the agency as a high-profile category of "new molecular entities." The agency also took an average of 17.6 months to approve the experimental drugs, compared with only 12.6 months for the 35 it approved in 1999.
FDA passed five drugs in 2000 within six months or less. They include Abbott Laboratories Inc.'s anti-HIV treatment Kaletra and Pharmacia Corp.'s antibiotic Zyvox. Almost a dozen others, however, took more than 18 months, including American Home Products Corp.'s ulcer medicine Protonix and Novartis AG's Exelon for treatment of Alzheimer's disease.
In 1993, the agency took an average of 26.5 months to approve new drugs. But it steadily picked up the pace, halving approval times by 1999. A 1992 law does not impose drug-approval deadlines on the FDA, but it obliges the agency to complete its review within one year—or within six months for "priority-review'' drugs that have a new method of action or have clear advantages over existing therapies.
CDER director Janet Woodcock says the agency met all of the goals imposed by the 1992 law last year, including a review of half of all original new drug applications within 10 months.
"We have not slowed down reviews. We exceeded all our user fee goals for review this year," she says, noting figures are available on the agency's Web site. "If there are fewer approvals, and we are reviewing just as fast, either there were fewer products submitted, or we turned more down."
Reviews, she says, are reactive, and CDER has no control over what drug developers submit. "If we send them an approvable letter and they take a year to respond, then that is a matter of their priorities and will therefore prolong the time it takes that product to get approved by a whole year," Woodcock says. "Approval times reflect how long the process took, both the number of cycles and the company time."
CDER has also been tied up with recalling drugs, including Pfizer Inc.'s Trovan antibiotic, because of safety concerns. Safety and efficacy are two priorities that reverberate throughout much of the FDA, which could account for the delays in approvals.
Silver lining
It is through clinical trials and approvals where the silver lining of genomic technology can be seen.
CDER's Dr. Sistare and Viragen's Rothberg agree that on one hand, use of genomic technologies could cost big bucks to implement and regulate. However, they are optimistic that the emerging technology could yield a tremendous cost savings and give drug makers a venue for tailor-making therapies based on the human genetic code.
"We have just unlocked the door," Rothberg says. "We are at the nursery school level right now, and we believe the scope of this will be so complex that's it's going to be an incredible challenge to meet the demand for these products."
Although it is in the early stages, Rothberg and Dr. Sistare believe a very important equation has been identified, a link to better, safer and more effective therapies at a lower cost for larger populations in a shorter amount of time.
"The promise of genomics will mean less failure and more efficient development of drugs," Dr. Sistare adds. "It is going to maximize the learning that occurs in clinical trials, in terms of better identifying patients and drugs that can serve them."