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Can Your Work Contribute to Biodefense?

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Three years ago, Luis Villarreal, a virologist at the University of California, Irvine, came up with the idea of using in vitro gene activation by PCR to study the proteome of a virus in hopes of developing a high-throughput platform for identifying new vaccines. Villarreal and his collaborators at the university saw infectious disease as their primary target, but after the anthrax attacks in late 2001 they realized that their work might just as easily apply to tackling bioterror agents. When the National Institute for Allergies and Infectious Diseases began soliciting research proposals for its new biodefense research initiative in the spring of 2002, Villarreal saw his chance to help the nation address a security challenge while perfecting his platform for future investigations. NIAID gave high scores to his proposal for applying the platform to the Francisella tularensis proteome (the bacterium that causes tularemia), although the money is currently bogged down in budget battles.

Villarreal is not the only scientist to realize that biodefense is fertile soil for growing genomic and proteomic research projects. With a projected biodefense research budget of $1.63 billion for fiscal year 2004, NIAID is just one of a handful of government agencies funding work in this area. Considering the mind-boggling sizes of the agencies’ biodefense budgets, opportunities for big biology are endless: The new Department of Homeland Security’s Science and Technology Directorate has requested $803 million for biodefense research in FY 2004; the Defense Advanced Research Projects Agency says it will allocate $137.3 million next year; the Centers for Disease Control, together with its National Center for Infectious Diseases, will have $8 million for 2004; and the armed forces control additional monies for biodefense research, although officials declined to release exact figures.

Suddenly, scientists eager to tap into this largesse are striving to show how their genomics and proteomics expertise is suited for describing the basic biology behind pathogens, developing tools to detect their presence in infected patients and in the environment, and for developing biodefense-related diagnostics and therapeutics.

And funding agency officials seem just as eager to hear from them. Through a combination of top-down directives and proposal-driven initiatives, NIAID, DHS, DARPA, and DOD’s armed forces divisions are all looking to arm themselves with a wide array of biological research tools — including those associated with big biology. The applications of genomics span the traditional paradigms of basic and applied research, says NIAID Deputy Director John La Montagne.

The government’s biodefense initiatives could be a boon to the genomics industry in more ways than one. The same way DARPA spun off technologies useful for civilian applications, such as the Internet, the biodefense effort is sure to lead to genomic and proteomic innovations applicable even to diseases unlikely to be used as bioweapons. What’s more, by providing a guaranteed customer — the federal government — for any therapeutics or vaccines against known bioterror agents, biodefense spending could help stabilize the biotechnology industry.

What remains to be seen is how well the various federal agencies will coordinate their potentially overlapping assignments. Divvying up $1.5 billion through NIAID alone will be hard enough. Synchronizing the priorities of multiple agencies as the field evolves could become a logistical nightmare.

How long biodefense will remain a critical priority is also in question. Despite the sense of urgency inspired by the 2001 anthrax attacks, many researchers who have already been promised funding are still waiting for checks to arrive. NIAID’s La Montagne says, “The fact is when you deal with a large increase like we’ve had [in our budget], it’s not something that you do overnight.”

Meantime, other agencies with new-found biodefense budgets are still defining their missions and communicating those goals to each other and their constituencies. Figuring out how each of these agencies plans to disburse its funds requires some degree of speculation, but through interviews with officials and a rigorous rummage through publicly available documentation, GT is able to offer a look at the biodefense funding landscape and a projection of the consequences for genomics and proteomics research.

NIAID

NIAID is the logical starting point for genomics and proteomics researchers looking to participate in biodefense. The agency has a mission to address infectious disease through understanding basic biology and developing new medical tools, combined with the largest biomedical budget directed toward biodefense. NIAID has already taken a leading role in sequencing the genomes of major pathogens such as anthrax and smallpox, and it has said in a published research strategy that genomic and proteomic data can contribute greatly to advancing its six areas of emphasis: microbial biology, host response, vaccines, therapeutics, diagnostics, and research resources.

NIAID is giving particular weight to proteomics and genomics as tools to gain insight into basic microbe biology. In addition to sequencing genomes, the institute has plans to develop central bioinformatics resources and tools for researchers to make rapid use of genomic information, and to apply genomics and proteomics to unraveling the virulence mechanisms of pathogens to identify potential new targets for vaccines, therapeutics, and diagnostics.

Genome- and proteome-based analyses should also come into play in the institute’s initiatives to investigate the mechanisms of host response to infection. Specifically, NIAID hopes to map the protective epitopes for several classes of pathogens using computational methods, immunochemistry, and structural biology, as well as genomics and proteomics. Diagnostic tools, too, should profit from efforts to identify patterns of gene and protein expression useful as signatures for pinpointing an infected patient.

NIAID provides a variety of mechanisms for researchers to gain access to its projected $1.63 billion fiscal year 2004 budget, including RO1- and SBIR-type grants and contracts. In addition, the institute is planning to fund six to 12 Regional Centers of Excellence for Bioterrorism and Emerging Diseases Research with grants of $65 million, which could provide additional avenues for researchers at smaller universities to engage their labs in larger biodefense projects, says David Gorenstein, a structural biologist at the University of Texas Medical Branch in Galveston who is the PI on a proposal representing several southern states.

CDC/NCID

CDC was a late arrival to the biodefense scene. But now, through a joint review process with NIAID, the agency is looking to direct extramural funding toward environmental detection systems for bioterror agents, technology for decontaminating areas exposed to pathogens, and research to determine the most effective means of delivering vaccines to the population. Among the latter would be studies comparing the efficacy of monoclonal antibody-based vaccines to other “therapeutic modalities,” says Bill Jarvis, director of extramural research for the CDC and NCID.

Jarvis and his colleagues are interested in new PCR-based pathogen detection tests, as well as methods for distinguishing between live and dead organisms, since in some cases only a live organism is virulent. The CDC is currently soliciting a broad range of proposals, Jarvis adds, as he and fellow administrators are still discussing which projects to tackle in-house and which to fund extra-murally. “Initially we wanted to be as broad as possible,” he says, while the CDC and NIAID work through which proposals match most closely with the two agencies’ respective missions. The first batch, received by the agencies last February, is still under review.

DARPA

Compared to other federal agenices, the Defense Advanced Research Projects Agency is an old hand at biodefense research. DARPA’s biodefense budget is actually dropping between fiscal years 2003 and 2004, from $163 million to a projected $137.3 million. But the agency will continue to fund high-risk research with long-term potential for biodefense application. Currently the agency funds biodefense projects in such areas as “on the fly” synthesis of oligonucleotides for use in pathogen detection systems, the genetic manipulation of organisms to create remote sentinels for the presence of pathogens, design of advanced diagnostics, and sequencing of pathogenic organisms such as Brucella suis and Franciscella tularensis. Janet Walker, a DARPA spokeswoman, says the agency contracts out all its research to university, nonprofit, and industrial researchers after defining specific project goals and soliciting proposals.

DOD

Through the Medical Biological Defense Research Program, the Department of Defense is directing funds both internally and with contract awards toward developing vaccines or other prophylactic treatments to prevent infection from potential bioterror agents such as anthrax, plague, Ebola, and pox viruses. The program is also funding tools for identifying and diagnosing patients infected with these agents, as well as medical treatments for curing those infected, says Carol Linden of the US Army Medical Research and Materiel Command in Fort Detrick, Md.

More specifically, Linden says her organization’s vaccine development efforts include host-pathogen interaction studies that target the pathogen’s mechanism of action and interactions with the body’s immune system, safer means of passive immunization such as human monoclonal antibodies or modified antibodies, and improved methods for vaccine delivery including nucleic acid vaccines and sustained-release formulations.

With respect to drug development, Linden says her branch of the US Army is employing strategies that include computational chemistry, combinatorial organic synthesis, high-throughput in vitro screening, and x-ray analysis of ligand-toxin co-crystals. In addition, the Army is using pharmacokinetics and mechanism-of-action studies to learn more about how potential therapeutics affect the body’s ability to respond to a pathogen. As for identification and diagnosis, Linden’s organization is supporting studies of assay systems that employ probes such as antibodies, metabolites, synthetic antigens, or nucleic acids. All of this work, Linden adds, is coordinated in close contact with NIAID efforts to avoid duplication.

In addition to biomedical research, the Army also has use for genomics-based tools in its programs for detecting the presence of pathogens in the environment, run primarily through the US Army Soldier Biological and Chemical Command laboratory at Edgewood Chemical and Biological Center.

James Valdes, the chief scientist at Edgewood, says his laboratory is testing the performance of various gene-based assays for detecting pathogens in the field, as well as using gene expression profiling experiments to look for expression patterns unique to low-level exposure to an agent. “For a terrorist suspect, there could be a gene expression profile that shows the suspect has been handling a terrorist agent,” he says. Although Valdes declined to state the size of his budget, he says Edgewood performs most of its biodefense work in-house, with extramural funding directed toward contract awards and SBIR-type research grants.

DHS

Charged with domestic security against terrorism, the Department of Homeland Security serves as a large umbrella organization for a wide range of research and technology programs aimed at detecting and combating terrorist attacks. Under the aegis of the Technical Support Working Group, DHS is sponsoring the development of new systems for detecting biological weapons in food and water, decontaminating individuals exposed to a biological weapons attack, detecting improvised biological weapons, and devising improved techniques for recovering DNA evidence from crime scenes.

Although the DHS public affairs office did not respond to repeated requests for information by press time, the department’s science and technology responsibilities range from basic science to advanced technology to actually deploying the new technologies in the field, says Sandra McCutchen-Maloney, a proteomics researcher at Lawrence Livermore Laboratory who attended a DHS workshop in early June to review the status of the many research projects whose funding the department now controls. “It’s been a whirlwind for them, but they really seem to have their act together,” she says. “They have a clear vision of where they’re going, and what’s needed. It’s really impressive; I was expecting a little more chaos.”

Looking Ahead

Despite these auspicious beginnings, program administrators acknowledge that the task of coordinating non-duplicative initiatives will be daunting, and that conflicting directives from higher-ups in the administration could complicate an already intricate budget-balancing process. Multi-agency workshops, collaborations across agencies, and demonstration projects all help researchers to avoid overlapping their colleagues’ efforts, says Pat Fitch, the program leader for chemical and biological national security at Lawrence Livermore National Laboratory. But there are instances when two groups “are each trying to solve the same problem, and 80 percent of what they’re doing is very similar and 20 percent is different,” he says. “How do you get another 60 percent worth of productivity with all this overlap? The short answer would have been DHS has got to serve a big role in that.”

Luis Villarreal, the UC Irvine virologist, says he’s worried that confusion over priorities at the government funding agencies — in his case NIAID — could create problems for researchers who expected funding but were then told to wait while administrators rejigger their budgets. Most recently, he says, the administration’s proposed budget dictated that NIAID should spend $250 million to purchase a given quantity of anthrax vaccine as part of the President’s Project Bioshield, which may have thrown a wrench in the institute’s schedule for paying out its approved grants.

La Montagne at NIAID says that the anthrax vaccine purchase could have contributed to delays, but says complicated questions over funding take time to resolve. “This is something for the long term, this is not something that’s going to go away tomorrow. The best way to deal with that is to build a solid, responsible research enterprise dealing with these problems in an effective manner.”

 

At Lawrence Livermore, Laying the Groundwork for Vaccines With Genomics

As genomics and proteomics researchers at Lawrence Livermore National Laboratory, Andrew Quong and Sandra McCutchen-Maloney are at the right place to apply their work to biodefense. The laboratory has a large appropriation formerly administered by the Department of Energy, now by the Department of Homeland Security, to address questions involving both basic biology and advanced technology for detection systems.

Quong’s work centers around gene expression studies of Yersinia pestis, the bacterium responsible for the plague, using both mathematical modeling and bench experiments to investigate relationships between the organism’s genes. With Y. pestis as his model system, Quong is trying to understand basic virulence mechanisms by correlating the organism’s gene expression profile with its phenotype under various conditions. “It is a good model system because there are close relatives to it,” Quong says. “Besides pestis there’s pseudotubercolosis and enterocolitica, so you have a nice way of doing comparative analysis with some very close near neighbors.”

McCutchen-Maloney uses multiplexed 2D gel electrophoresis combined with mass spectrometry to study changes in protein expression in cells exposed to Y. pestis as a means of identifying markers with potential as diagnostics or therapeutic drug targets. “It’s a new twist to an old trick,” she says. “We look at multiple samples on one gel using fluorescent labels, and the fluorescent label actually improves the detection by an order of magnitude from traditional 2D electrophoresis.”

LLNL is placing heavy emphasis on genomics and proteomics as tools for biodefense partly because Pat Fitch, the program leader for chemical and biological national security at the lab, believes the tools will be important in laying the groundwork for new vaccines. Because anthrax and other potential biowarfare agents are so lethal and so rare, it will be difficult for researchers to construct meaningful clinical trials, forcing vaccine developers to rely heavily on more basic biological information obtained from genomics and proteomics studies, he says. Furthermore, Fitch hopes NIAID’s large investment will “leapfrog our understanding of how infectious diseases work and how our immune system works” to where researchers can use a small panel of markers to identify large classes of virulent organisms. “Things like that, we’re not going to be able to do unless we get the next generation of genomics and proteomics tools.”

—JSM

 

Agilent Technologies’ Mobile Laboratory: A Vehicle for New Detection Technology

Designed initially by Army researchers at the Edgewood Chemical and Biological Center, part of the US Army Soldier Chemical and Biological Command, the mobile laboratory is now marketed by Agilent Technologies to the military, local governments, and private-sector companies. Last year, Agilent donated a mobile lab to the New York City Police Department, the first to be employed for detection of chemical and biological warfare agents in the civilian theater.

Fabricated by ENG Mobile of Concord, Calif., the mobile lab has two compartments — the rear for hazardous sample submission and the front for sample analysis. Each compartment has separate HVAC and air exchange, air filtration and pressure settings. Purified Microenvironments of Miami, Fla., manufactures the air-handling equipment, developed with the technical assistance of US Army scientists.

To detect and identify biological pathogens down to the specific strain of bacterium, MIDI of Newark, Del., provides a system that employs the fatty acid methyl ester signature of each microorganism to identify all major bacterial biowarfare agents (such as anthrax and plague) as well as many other pathogens. The system includes an Agilent 6850 gas chromatograph, ChemStation system and MIDI Sherlock software. In addition, the lab is outfitted with an Agilent 2100 Bioanalyzer that relies on Caliper Technologies’ microfluidics chips to sort and measure quantities of DNA, RNA, and proteins from a sample prior to identification.

—JSM

 

Raising the Limits of Detection with New Microarray and Mass Spec Technology

Lest one think Agilent Technologies’ version of a mobile laboratory will be the only game in town for the indefinite future, DARPA-funded researchers are hard at work developing the next generation of gene- and protein-based devices for detecting biowarfare agents. At Johns Hopkins University’s Applied Physics Laboratory in Baltimore, Joany Jackman and Tim Cornish are designing new microarray and mass spectrometry systems that promise to make biowarfare agent detection more rapid and more accurate.

Unlike other gene-based microarray systems, Jackman’s gel-based microarray detects ribosomal RNA, allowing her group to avoid both normalizing for gene expression and PCR amplification, since the organism has already transcribed multiple copies of the sequence, she says. In addition, Jackman’s microarray uses many tests for the same expressed gene, raising the confidence of the system’s detection.

Meanwhile, Cornish and his collaborators at APL are busy testing prototype versions of a miniature MALDI-TOF mass spectrometer for detecting biowarfare agents in the field. The system collects an aerosol sample and looks for coat proteins or other biomarkers indicative of the pathogen, without requiring sophisticated sample work-up, Cornish says. The advantage over conventional PCR-based gene tests, which can take several hours to complete, he adds, is that the mass spectrometry approach provides the basis for real-time detection. Furthermore, mass spectrometry is more amenable to automation, he says. Cornish’s group has completed testing the second-generation version of the instrument, and is preparing a third-generation prototype.

—JSM

 

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