The program will require the talents of engineers, physicists, chemists, mathematicians, computer scientists, and molecular biologists. Many industries, including biotechnology, microelectronics, software, instrumentation, pharmaceutical, and fine chemicals, will be called upon to participate in this critical national effort. Through these collaborations, technological advances in microchemistry, micromachining, microfluidics, separation technologies, detection systems, microelectronics, and information technology will be efficiently integrated.

The technical goal of the program is to develop cost-effective methods to determine, analyze, and store DNA sequences for a wide variety of diagnostic applications, ranging from health care to agriculture to the environment. These systems will be automated, miniaturized whenever possible, high-throughput, accurate, low-cost, and user-friendly.

In medical diagnostic applications, for example, a suitable system might begin with the injection of a biological sample into a cassette, which would automatically be positioned in a reader. All the steps of the analysis would be performed automatically and accurately, and the results displayed on a computer screen and immediately transferred to the patient's computerized record. For environmental or agricultural uses, readily transportable, miniaturized, hand-held devices are envisioned. Sequencing instruments for studying the effects of environmental mutagens require a very high degree of sensitivity since the goals will include searching for rare genetic changes in cell populations.

The business goal of the program is to support the development of a new and very large, potential market for DNA-diagnostic systems. Recent advances in DNA technology have set the stage for development of systems that will have a wide variety of commercial applications. Successful accomplishment of these goals would create new opportunities in many fields, including health care, agriculture, veterinary medicine, environmental monitoring, and personal identification.

The program should enable industry to deliver DNA diagnostics to a variety of industrial sectors at 1/10 to 1/100 the current price. It should also enable the industry to reduce the cost of DNA sequencing and make DNA sequencing apparatus available at significantly less than the current cost.

DNA is the universal code for all biological organisms. The availability of technology which provides cost-efficient, sequence-based analysis of that code will affect virtually all industries that currently rely upon or service biological organisms. Therefore, this program will not only provide for cost savings in existing markets such as diagnostics, drug discovery, forensics, and infectious-agent identification, but will allow for the creation and expansion of new markets and applications.

In the long term, DNA sequence/diagnostic analysis technologies are likely to yield increased activity and greater efficiencies in such areas as:

• health care;

• forensics and personnel identification;

• biomedical research, including pure and applied biology;

• environmental monitoring;

• toxicology;

• drug discovery and design;

• biomass diversity, assessment, maintenance,

• bioremediation;

• infectious agent monitoring;

• quality control in the food industry;

• industrial processing, including the monitoring of biological processes; and

• animal husbandry and agriculture, including assessing and maintaining diversity, improving yield, monitoring disease resistance, assessing nutritional value, and targeting biocontrol.

Together with the information that will be forthcoming from the Human Genome Project and other genome efforts, this technology can aid in increasing the use and decreasing the cost of DNA diagnostics for preventive screening, symptomatic and presymptomatic testing; create a market for cost-effective therapeutic monitoring of patients; and create a market for monitoring of endogenous gene expression to aid in informed diagnosis, the combination of which should yield more individually-tailored rational therapy regimens.

Diagnostic testing is one of the fastest growing applications of biotechnology for health care, growing at a rate of 15 to 20% annually. Economic and market data are based on estimates obtained from various industrial sources. The total U.S. market for in-vitro diagnostics is estimated at $5 billion in 1992 and is comprised of clinical chemistry ($2.2 billion), immunoassay ($1.6 billion), hematology ($0.7 billion), and DNA probes ($0.06 billion). The projected markets for health care diagnostics is expected to grow to over $20 billion dollars by 2000. The DNA based in-vitro diagnostics component of this is market is currently very small but is expected to grow substantially.

If DNA diagnostic technologies were available that were one to two orders of magnitude faster and an order of magnitude cheaper (current testing kits for monogenic diseases range from $125 for cystic fibrosis to $350 for Duchenne/Becker muscular dystrophy), it could substantially expand the market as well as reduce the necessity for other less desirable assay technologies.

DNA diagnostic tests for agriculture could supplant the presently used methodologies to identify plants with special characteristics such as hardiness, diseases resistance, etc. The use of these tests will become widespread if the cost of analysis is significantly reduced. In a similar way, use of this technology will become widespread in animal genetics, plant and animal design, husbandry and the treatment of disease. Market penetration will very much depend on the costs of these tests. For example, a reduction of the costs to ten dollars per test will lead to a use of these in the control of the germ lines in artificial insemination of cattle. Furthermore, the reduction of the costs of doing DNA analysis will widen the market for these instruments and diagnostics. Chemical process technologies will be affected by this program. It is expected that inexpensive DNA diagnostics will be used to monitor bioprocessing for the production of specialty and commodity chemicals. At present, there exist no apparent barriers to the commercialization of the resulting products from research applications.

Other applications related to health care and the agribusiness are likely to require approval by the Food and Drug Administration as well as more thorough optimization of conditions, definition of standards, and evaluation of error rates. These issues should be addressed in applications for awards. There is precedent for the use of biological or DNA-based technologies for most of the applications listed above and therefore, the potential to achieve commercialization has been demonstrated, although optimization of the process for individual cases will still need to occur.

Some recent studies of DNA technologies are abstracted below.

• Best Practices: Benchmarking & Consulting, LLC., "DNA-Based Technologies Forecast". 1997.

Best Practices, LLC. Executed a benchmarking methodology that would help leading companies and academic laboratories better evaluate the future impact of DNA-technologies on diagnostics, drug discovery, disease management, and other related subjects. A major focus of the report was likely scenarios for DNA-based diagnostics in the next ten years. The purpose of these scenarios is to enable laboratories to make more informed strategic decisions about future plans for research, investment, and acquisitions or partnerships in the diagnostics arena.

Best Practices gathered the information through an expert opinion benchmarking exchange among leading scientists in 45 DNA laboratories. The results predict that PCR will have the greatest market impact by year 2007 with DNA chips/arrays and gene expression not far behind.

Both academia and industry were overwhelmingly positive about the future potential of DNA-based technologies. The benchmarking participants assessed a higher than 70% (weighted average) probability that DNA-based technologies would be the dominant platform for discovering and developing both diagnostics and pharmaceutical products. They also felt that there is a greater than average chance DNA technologies would play an integral role in disease management and selecting treatment regimes.

• Regalado, Antonio. Start-Up, "The DNA-Chip In Diagnostics", vol. 1, no. 2, Sept. 1996, p. 18.

This article provides an overview of how DNA-probe arrays and their use in diagnostics has moved from concept to the marketplace over the last five years. It also assesses the opportunities of DNA-chip technology in diagnostics and how some of these opportunities such as point-of-care testing and infectious disease monitoring may be put on hold because of ethical, competitive and technological reasons.

Regaldo highlights several young companies funded by the "Tools for DNA Diagnostics" program at NIST's Advanced Technology Program (ATP) to develop integrated and miniaturized systems for DNA diagnostics. These companies are developing some of the hottest technologies and strongest patent positions.

• Regaldo, Antonio. Start-Up, "DNA-Chips In Genomics", vol. 1, no. 3, Oct. 1996, p. 24.

This article discusses the opportunities of DNA-chips in genomics and how DNA-chip companies are being attracted to this area. It also highlights the work of ATP awardees participating in the "Tools for DNA Diagnostics" program and the opportunities they have to become major players in this industry.

The short term market opportunities for many of these young companies may come from genomic collaborations rather than from diagnostics.

• The National Cancer Institute, "The Nation's Investment in Cancer Research", A Budget Proposal For Fiscal Years 1997/1998.

NCI is launching a large scale effort in cancer genomics. By working closely with scientists, educators and community leaders, NCI has identified five programs that need immediate investment. These programs are:

• Cancer Genetics

• Preclinical Models of Cancer

• Detection Technologies

• Developmental Diagnostics

• Investigator-Initiated Research

This effort is in response to recent advances such as rapid identification of new cancer genes, to recent breakthroughs in the ability to develop preclinical models of cancer, or to looming social responsibilities such as the issues raised by genetic testing.

NCI's goal is to assure that the Nation's cancer research resources are used as effectively as possible.

The following discussion addresses some of the technical areas eligible for proposals in the Tools for DNA Diagnostics Program together with some examples.

At present, there exist three approaches to DNA analysis that appear to be the most promising for automated DNA diagnostics applications: serial sequence analysis, hybridization analysis, and amplification-based analysis. Previous government investments in basic research, including the Human Genome Project, have supported the acquisition of preliminary data to demonstrate the proof of principle of these approaches. However, these programs, including the Human Genome Project, will not support the development of these techniques for use in diagnostic applications.

The utility of DNA diagnostics already is apparent, even with only a small proportion of the human genome sequenced. The application of DNA diagnostics will greatly expand as more of the human genome is sequenced. The use of any of the three approaches listed for diagnostic applications will require miniaturization, parallelization, and automation of current technology. Independent of the approach, improvements will need to be made in the areas of sample preparation, assay technology, detection systems, integration, and data management and analysis. Specific improvements in each of these areas will depend upon the specific approach and the ultimate diagnostic application of the technology. Beyond this, however, many of the technological improvements that are required are sufficiently basic and enabling that the improvements will benefit industries and markets beyond DNA diagnostics.

In the areas of sample preparation and assay, it is clear that miniaturization is key. To reduce the size of samples by a factor of 10 or greater, barriers in microfluidics, micromachining, robotics, microchemistry, nucleic acid chemistry, and surface chemistry must be overcome. To implement miniaturized protocols accurately and efficiently, substantial automation of the process will be required. In the development of miniaturized systems, it is essential that the system can be adapted for high levels of parallelization.

Miniaturization

Miniaturization poses significant technological risks. Currently, there exists no universally accepted precedent for the handling, replication, amplification, or cloning of DNA in nanoliter volumes. Due to the size and charge of the DNA molecule, and the relative instability of many of the enzymes involved in the sample preparation processes, nanoliter and less volumes may pose substantial challenges. In addition, interactions of the biological molecules with the surfaces of the reaction chambers must be minimized. For some methodologies, it is not clear what the optimal sample will be, so substantial improvement in DNA fragmentation technologies or DNA cloning vectors may be required for the ultimate efficient application to diagnostics. Improvements in any of these areas are likely to be of value to other non-DNA based diagnostic applications such as antibody screening protocols and enzyme-based diagnostics, because miniaturized robotic or micro-electro mechanical systems developed for DNA could be modified to be used for these purposes.

Assay

Assay capabilities improvements for DNA diagnostics are likely to be somewhat dependent upon the specific approach. For instance, in serial sequence DNA analysis application of capillary gel electrophoresis, ultrathin gels, and microchannel arrays to the separation step will increase the speed and parallelization of DNA analysis by a least one order of magnitude. New efforts that are being undertaken to apply microfabrication and micro-electro mechanical systems technology to the separation step could afford an increase in the speed of two to three orders of magnitude. All of these approaches require the development of improved separation matrices. In addition, improved capillary fabrication or microfabrication will be required and fluid handling and surface chemistry issues addressed.

Assay capabilities for hybridization-based DNA analysis also will require overcoming a variety of technical barriers. Improved technologies for the production of low cost oligonucleotide arrays will need to be developed. This requires new approaches to oligonucleotide synthesis, improved surface chemistry for oligonucleotide attachment, and microfabrication improvements ultilizing many substrates including silicon, glass and polymers. Optimization of the biochemistry involved in the actual hybridization is also required and may be specific for the detection format. Automation to allow for the processing of many parallel units is also be required.

For amplification-based DNA analysis, there are two possibilities for the assay step, one of which includes a separation step similar to that used for serial sequence analysis, and therefore would require overcoming similar technical barriers to those listed above. An alternative assay would involve enzymatic detection similar to current antibody assays, which could possibly allow for future "dip stick" formats for particular diagnostic applications. This assay would require the development of biochemistry for the attachment of appropriate enzymatic substrates to DNA and optimization of surface chemistry to promote specific sample binding and prevent anomalous sample binding. As with other approaches, automation to allow for the processing of many parallel units will be required.

Detection

The detection step of each of the three DNA analysis approaches will require both improved reporter groups and improved detector technology. With the proposed miniaturization described above, detection will have to be increasingly more sensitive. In addition, new detection technologies could allow for increased multiplicity of samples per unit of the assay step. A limited number of fluorescent, energy transfer, or infrared dyes constitute the most commonly used current reporter groups. Additional dyes are being sought, but other reporters such as electrophores and heavy metals are also being developed. Improved chemistry for the production of these reporters and the attachment of the reporters to DNA will be required. The sensitivity of the appropriate detectors for each of these reporters will have to be increased. The development of more sensitive charge coupled device (CCD) arrays and photodiode arrays may also be required. Some approaches to detection that do not rely on reporter groups include measurement of changes in impedance following hybridization and measurement of electron conduction along double stranded DNA. Optimization of such to allow parallelization and high throughput approaches requires development of improved micromachining and microelectronic capabilities.

Data processing

With the expected improvement in diagnostic capability, better systems will need to be made available to handle the increased information output. This will include better data management tools, and better analysis software. It is likely that individual software packages will be required for each type of application to make the systems user friendly. In addition, improvements in systems integration will be necessary for all applications, such that the interface between components of the individual systems is transparent to the user.

Integration

The ultimate challenge of all the areas described above is to integrate the steps outlined from sample preparation to analysis in an economic way so that DNA analysis can be performed substantially quicker for orders of magnitude less cost than existing tests for simpler monogenic diseases. This involves thoughtful integration in the preliminary design of the methodology to be developed.

Proposals are welcome in all of these areas as well as any other approaches that address the fundamental technical issues described here. This program will not accept proposals which differ in content and focus from the scope discussed above. Proposed projects should focus on developing enabling technologies; the ATP will not fund product development. Examples of proposals that will not be accepted include studies to design, develop or target drugs, agrochemicals, herbicides, etc. based on genomic information, studies involving gene discovery and the de novo sequencing of any DNA fragment or genome. Proposals that are primarily directed towards the development of lasers are also not included since laser development is already being supported in various ATP projects and programs.

This competition, the third, will focus on the development of basic technologies necessary for driving the costs of DNA analysis down to the target levels. Thirteen awards, ranging from less than $2M to over $30M were made in the first competition, held in 1994. Seven awards ranging from less than $2M to $7M were made in the second competition held in 1995. Additional awards have also been made in the 1992, 1996, and 1997 general competions. Recipients include both large and small companies, startups, and both joint ventures and single applicants. The funded proposals represent diverse technologies varying from electrophoresis to hybridization to PCR methodologies.

The strong industry commitment to a program in the area of improved technologies for sequenced-based DNA analysis is clear from the number and quality of white paper submissions (this area was the subject of more white-paper proposals than any other biotechnology-related topic.) and to the quality of the proposals submitted to the competitions. The development of improved technologies for sequenced-based DNA analysis involves a diverse group of both large and small industries with varied expertise and interests in areas including, but not limited to, biological sample preparation and molecular biology; microfabrication; surface chemistry; separation sciences; nucleic acid chemistry; instrumentation development and engineering; detection technologies, in particular various forms of excitable wavelength spectroscopy; and information handling, data analysis, and systems integration. White papers defining programs related to this area were received from a broad spectrum of companies, including representatives of the pharmaceutical industry, drug development companies, molecular biologicals suppliers, instrumentation development firms, and software developers, as well as others. In addition, the contributors of white papers in this area were broadly distributed in size, ranging from large corporations to small start-up companies.

The real opportunity and enthusiasm for industry involvement in this area is also reflected by the significant level of industry participation in meetings of the scientific community related to this area such as the International Genome Sequencing and Analysis Conferences, as well as the appearance of national meetings directed toward industries interested in this area, such as "The Human Genome Project: Commercial Implications", "Genetic Screening and Diagnosis of Human Disease", "BioChips" and "Nanotechnology." But perhaps the most persuasive testimony to industry interest in this area is the number of start-up companies which have formed in response to the opportunities in this area.

There is a strong industry commitment to the development of DNA technologies for sequencing, diagnostics, and the development of drug and gene therapies. Large and small companies are involved in these activities. Several companies are involved in the development of the concepts and technology necessary for oligonucleotide array hybridization, capillary electrophoresis, and mass spectrometry. Laser development is being pursued by several companies for the development of cheaper, more efficient, solid-state lasers. Micromanufacturing technologies are being pursued for the manufacture of chips, capillaries and detector arrays.

The United States has achieved global leadership in basic biology and biotechnology in the world-wide biotechnology industry. This leadership is the result of significant past investments in biological research by both large and small companies and, particularly, government. Maintaining leadership will require identifying and nurturing the development of key technologies. Increasingly, the key technology in biotechnology is DNA analysis, given the broad range of commercial applications for diagnostics, therapeutics, bioprocessing, agriculture, forensics, and toxicology.

At present, the major contribution of the U.S. government to the general area of DNA analysis is through support of the Human Genome Project. This project, sponsored by both the National Institutes of Health (NIH) and the Department of Energy (DOE), supports basic research toward technology development in the area of large-scale genomic DNA sequencing, as well as the use of state-of-the-art technologies for sequencing selected small model organism genomes. The Human Genome Project, however, does not support the development of technologies for diagnostic applications, which will ultimately be quite different from the technologies used to acquire the sequence of the first human genome. Government support for development of DNA diagnostic technologies for health-related, forensic, agribusiness, and toxicological applications is currently small.

The results of basic research supported by the Human Genome Project on new technologies for large-scale genomic DNA sequencing will provide a scientific foundation for the development of technologies for DNA diagnostics. In addition, the sequence information that has been obtained from the human and other genomes, and the complete human genome sequence that is the final goal of the Human Genome Project, will provide the biological foundation for DNA diagnostics. The proposed ATP program on DNA diagnostics can leverage the existing government investments in these areas to achieve the aim of low-cost DNA diagnostic technologies. These technologies coupled with the information to be obtained through the Human Genome Project can allow for the explosion of applications for DNA diagnostics in the health care industry, and the ensuing technology can have far reaching impact on creating markets in the agricultural and livestock industries.

The NIH and DOE Human Genome Program and NCI have agreed to provide technical staff to support the ATP program and ensure appropriate coordination. Although technological and marketing goals outlined in this program are believed to be achievable within the next decade, industry has been hesitant to invest sufficient sums to drive this area without government support. At present, there are various competing methodologies and no guarantee exists that any one technology will be the choice for the long-term applications. Therefore, significant risk is associated with introducing specific technologies in a competitive manner. For individual companies the associated technical and marketing risks, in combination with the high costs of technology development, lead to a reluctance to pursue these opportunities with vigor. While the risk of investing can be high for individual companies, for our nation the greatest risk is to not encourage this technological area and to lose out on the opportunity to create new DNA diagnostic markets, reduce health care costs, and sacrifice our lead in the field of biotechnology to fierce international competition.

The ATP offers tremendous opportunity for both vertical and horizontal integration of industries. This is exemplified by the wide variety of current ATP single applicant awards in this program area and several joint ventures. Without ATP funding, this technology development effort would not be happening. Small businesses lack the funding to undertake risky projects that require many different types of technological advances. Larger businesses also have difficulties in justifying the significant investments required to develop high-risk, high-payoff technologies through to commercialization given the long term nature of the investment. Furthermore, even large companies do not have all the expertise needed for such multidisciplinary projects.

Through the ATP DNA diagnostics program, the resources and new developments resulting from small companies doing basic research and developing and replicating limited numbers of prototypes can be combined and appropriately leveraged by larger industrial partners, eventually leading to integrated, commercial products. The potential investment of ATP in this area could serve to promote these interactions and provide financial incentives for industry collaboration.

The ATP Tools for DNA Diagnostics program will greatly enhance the opportunity for companies to form highly competitive consortia, allowing them to maximize the chance for technological success while sharing the burden of potential generic technology development risks. No other government program is well suited to meeting this challenge in an aggressive and timely manner. The long-term potential is to accelerate the building of U.S. DNA diagnostic technology industrial base that will promote economic growth and the quality of life in the 21st century.

Date Created: December 1997
Last Update: April 8, 2005