The Fun Guy's Journal: Scientists and Salesmen: the New Ownership of Biotech

(Why post something this long? Because I can.)

Scientists and Salesmen: the New Ownership of Biotech

Brendan A. Niemira

Abstract

The biotech revolution grew out of private enterprise and publicly funded effort, sometimes competing, sometimes cooperating, but always intertwined. With new applications of biotechnology maturing ever more rapidly in the global marketplace, efforts to use the flood of information and novel techniques to balance profit and the public good are also emerging. This essay will review some of the current advances in biotech, primarily from a food standpoint, and some of the ways these advances are being received by the public.

Introduction

        The phrase "Biotechnology Revolution", so much in vogue in the 1990's, isn't commonly used in 2005 to describe what is happening in the life sciences, but only because the revolution has become mainstream. Much like earlier quantum leaps of science and technology, such as the Industrial Revolution, the Green Revolution and the Personal Computer Revolution, these revolutions are ongoing, and continue to shape our world in ways the scientific pioneers could never have imagined. With every new advance, science becomes industry, the revolutionary becomes commonplace and novelties become necessities.

        The new technologies that continue to spring from these revolutions offer new efficiencies, new capabilities, or entirely new industries. In every case, from machine tools to hybrid plants to the Internet, early adopters of the various new technologies either became fabulously wealthy or went bankrupt, or perhaps both. The second wave of adopters, having learned from the experience of the early adopters, began the process of sorting the wheat from the chaff in order to best take advantage of the new technologies. In order to sort out the hypothetical from the possible, and then to make the possible into the feasible (and ultimately make the feasible into the profitable), they determined the limiting factors on the growth of their technologies. For each problem they faced, they asked themselves, "What knowledge are we lacking?", then set about acquiring that knowledge in a systematic way.

        If basic science is the exploration of terra incognita, then applied science is the systematic mapping of a coastline to find useful features of the landscape. Basic and applied science are two halves of the same coin, each relying on the other for sustenance, support and growth. Neither could long exist without the other, and the most productive fields are those in which each approach is given its due. Biotechnology, predominantly a basic science endeavor in its early years, is increasingly alloyed with applied science pursuits. Some regard this as a sign of maturity, others as a sign of corruption. This clash of perspectives has resulted in confrontations within the scientific community over the introduction of genetically modified crop plants, potential applications in livestock as well as in human medicine. These conflicting views shape the course of scientific advance. One of the most illustrative examples of this, well worth examination in some detail, was the effort to sequence the human genome, a heated competition that set the mold for all genomic research that was to follow.

Public vs. Private

        The Human Genome Project (HGP) was a large-scale scientific project to determine the sequence of all of the DNA in human cells, the entirety of the code that is makes humans what we are (11). The human genome was described in press accounts of the time as "the Book of Life". The hyperbole was excusable, since it was believed that once the human genome was fully known, then a cornucopia of benefits would flow almost immediately, from new drugs to cures for a myriad of diseases. This project, a monumental task, was first proposed in the early 1980's, and was considered so vast in scope and so costly to pursue that it would be beyond the ability of any private entity to undertake. An international consortium of government sponsored laboratories was created for the HGP, and a team of world renowned molecular biologists was recruited to oversee it. The great machinery of the HGP officially came into being in 1990, and although strands of DNA began to be sequenced and data started flowing within a few months, the project was expected to take many years to complete. This effort launched the field of genomics, a systematic approach to understanding how genes fit together on DNA, their relative positions and potential interactions. In 1993, the government funded scientists predicted that they would be able to complete the project in 10 or 12 years.

        One company, impatient for the riches that awaited the first to read the Book of Life, wasn't willing to wait. In 1998, a company called Celera, founded by a former NIH scientist, started its own project to sequence the human genome, using a novel DNA sequencing technique. The HGP's progress was relatively slow because it used the "gold standard" of sequencing. The scientists took relatively large pieces of DNA, laboriously sequenced them, and fitted them onto a map. An analogy would be to take pages out of a book and start reading each individual page, beginning at the first line, then attempting to reassemble the book based on prior knowledge about the plot. In contrast, Celera used the "shotgun" method, whereby large fragments of DNA were chopped into very small pieces using different kinds of enzymes. These small pieces could be sequenced very quickly using automated sequencing machines. In successive steps, the original DNA with the unknown sequence was cut in different places, such that the sequences of the resulting fragments overlapped. When the short sequences were known, a computer identified the overlaps in the sequences and used probabilities to stitch the small sequences into a coherent single sequence. To continue the book analogy, it would be the equivalent of tearing up a dozen copies of the book in a dozen different ways, then reading the couple of sentences on each piece and assembling a new, complete copy of the book from the overlapping fragments.

        Celera's approach gave results of lower quality than the gold standard method used by the publicly funded HGP. The probabilistic nature of the process meant that huge quantities of data would have to be generated and sifted before any meaningful sequences could be put together, the same portions of the genome would be sequenced over and over again and that it would result in lots of gaps throughout the assembled genome, which the shotgun technique was not well suited to fill in. However, it was much faster, much cheaper and could be scaled up very rapidly by adding more sequencing machines and more computers. During this period in the late 1990's, the rapidly increasing power of computers to accommodate the computational challenge improved the overall efficiency of the shotgun method. In short, while not as good as the gold standard, it was fully good enough to be of value to industry. Within months, Celera had sequenced a substantial portion of the human genome and was accelerating its pace of discovery, predicting that it would produce a complete genome map in 3 years, rather than the 7 called for on the HGP timetable. In this race between public and private effort, Celera enjoyed a crucial advantage. As a publicly funded project, the HGP released their DNA sequences to public databases as they learned them. Celera was able to incorporate the data into its own sequence, and take advantage of this information to fine-tune its own approach, filling in gaps and using the published sequences to improve the efficiency of its sequence assembly computers. Faced with the competition for the honor and prestige of completing the genome, the HGP consortium accelerated its pace, shortening the completion timeframe first from 7 years to 5 years, and then again to 3 years. The atmosphere turned acrimonious, and the leaders of the two groups traded public accusations regarding each other's motives. The two efforts, public and private, became a horse race much discussed in scientific circles. At stake, it seemed, was the very ownership of the human genome, and the expected flood of biotechnology miracles as well. The first group to produce a complete genome sequence would set a precedent for the Book of Life being, conceptually at least, either private property or a common resource.

        In the end, under considerable pressure, the competitors agreed to call it a tie. The public and private groups both agreed to release their maps simultaneously, although lingering animosity prompted them to eventually publish them in competing scientific journals. In June 2000, at a White House ceremony, both groups were hailed for their achievements and the race to sequence the human genome was announced to be officially over. Though the smiles at the ceremony were forced and the mutual congratulations were tepid at best, this episode stands as a classic example of public and private effort driving each other to greater achievement than either would have attained alone. The story is worth discussing because in it can be seen an archetype for how the different perspectives compete, and, by their competition, ultimately benefit society as a whole. Without the public effort, the sequencing of the human genome would never have been started; without the private effort, it would have taken at least twice as long and cost at least three times as much. Subsequent genome sequencing projects combined the approaches of both camps to combine the demonstrated speed and cost benefits of the shotgun approach with the high sequence quality of the directed sequence method. The best approach was not one or the other, but a combination of the two, a combination that never would have been attempted had the race never taken place. Ultimately, one of the most important results of the race for the human genome was the sequencing methods that were developed, rather than the sequences themselves.

        The information management and distribution systems set up to use the flood of human genomic data remain the infrastructure of the entire field. The Institute for Genomic Research (TIGR), the National Center for Biotechnology Research (NCBI), the European Molecular Biology Laboratory (EMBL), the DNA DataBank of Japan (DDBJ) and other genomics database site are used on a daily basis by scientists worldwide. These databases are publicly searchable, making the genomic data widely available; private companies charge additional fees for annotations, data analysis and other value-added services.

From Genes to Proteins...

        The great irony of the quest to sequence the human genome is that the Book of Life turns out to be a great deal more complicated that it seemed at the time. Obtaining the full DNA sequence of Homo sapiens was not the final step in unlocking how our cells and tissues work. When DNA was first understood to be the repository of genetic information, it was known that genes code for proteins, i.e. that the sequence of DNA in a gene determines which amino acids get strung together in which particular sequence to make a particular protein. Humans have roughly 20,000-25,000 genes, many fewer than the 80,000 to 120,000 genes which had been predicted. Humans have somewhat fewer genes than mice (30,000) and somewhat more than fruit flies (14,000). In comparison, the number of genes in various plants, such as rice (38,000), corn (60,000) and mustard (24,000), can match or exceed our own. In later years, a clearer understanding of the incredible complexity of how this system is regulated began to emerge.

        The U.S. government website for the Human Genome Project (5) quotes Winston Churchill to describe the state of understanding of the human genome in 2005, five years after it was fully sequenced: "Now is not the end. It is not even the beginning of the end. But it is, perhaps, the end of the beginning." Researchers refer to the current state of the art as the postgenomic era, or the era of functional genomics. It can be expressed prosaically: "Now that we know what genes we have, how do we determine what they do and how they are interacting?" It turns out that most of the activity of cells is determined, not primarily by the genes on the DNA, but by the interactions of the proteins and other cellular components. Depending on the complement of proteins in a cell, a given gene can make any one of several versions of its gene product, usually a protein, but sometimes a functional piece of RNA or a protein-RNA complex. Each of these different versions of the gene products has a different functionality. By combining, dividing, modifying each other, inhibiting or enhancing each others activities, the cell's proteins determine how a cell or tissue behaves. The collected entirety of genes and their interactions is what makes up a genome; by analogy, the constellation of proteins in a cell and their interactions is called a proteome. The study of this interaction of proteins is called proteomics. ... and from Genomics to Proteomics

        If the challenges of genomics are daunting, then those of proteomics are breathtaking. To be sure, the potential benefits are as miraculous as ever - understanding exactly why certain cells, tissues and entire organisms behave the way they do under any given set of circumstances is immeasurably valuable. The conceptual framework of observing how the entire proteome changes in response to stimuli has led to the establishment of other "omics" fields. These use genomic and proteomic techniques to focus specifically on particular issues within the larger question of the functioning of living cells, tissues and organisms. These "omics" fields include, investigations of, among others, the entire complement of transcription products ("transciptomics") how gene expression responds to specific food components ("nutrigenomics"), how cell constituents are broken down and recycled ("degradomics"), and how cells communicate using peptides ("peptidomics").

        One "omics" field which is the sine qua non for all other research is "economics", the investigation of which types of research projects get funded. Whether through private industry, university based grant funded research or publicly funded government agencies, the decision to fund one type of work or another, to buy equipment suited to one task or another, to hire either a virologist or an oncologist is driven by perceived potential for return on investment. As the technology behind genomics/proteomics research advances, the areas of inquiry to which it can be potentially applied multiply faster than finite resources can keep up. Everyone can theoretically benefit from genomics/proteomics; the question is, who gets to benefit first?

        The powerful new tools of analysis are developing at a furious pace, and serve to enhance the speed and accuracy with which organisms can be modified and manipulated. We have at our disposal a vastly expanded understanding of how life works, and an array of tools far more accurate and powerful than those of 50, 20, 10 or even 5 years ago. Genetically modified crop plants are commonplace, and increasing in adoption worldwide. The technology already exists to begin the creation of genetically modified livestock, albeit in a more limited way, but ethical and political concerns have stalled many efforts to directly alter the genetic makeup of higher vertebrates. Genetic modifications to humans are still hypothetical, but becoming more feasible every year. These technologies hold great promise for the development of new drugs and new genetics-based therapies, but many people are legitimately concerned that they could also be applied for trivial, unwholesome, or harmful purposes.

        It should strike no note of surprise to realize that the statements of the preceding paragraph could have been taken almost verbatim from ITEST's 1999 workshop, "The Genome: Plant, Animal, Human" (10). With years of additional progress in biotechnology and years of additional experience with the ecological, ethical and legal issues surrounding the creation and application of genetically modified organisms, has society's response to biotechnology developed as well? Has our situation really changed?

Yesterday's Tomorrow

          For a sense of perspective, the essayists for the current workshop were asked to review their essays presented at the 1999 ITEST workshop. My essay dealt with genetically modified plants, and, more generally, with the societal impact of genetically modified organisms (10). In that essay, I presented three key technological breakthroughs which were necessary for "the genetic revolution". The first was the identification of DNA as the carrier of genetic information, in the 1940's and 50's. The second was the development of sophisticated biochemical tools in the 1980's. These tools allowed scientists to manipulate DNA and included novel enzymes which cut DNA at specified points in the sequence (nucleases) or stitch DNA strands together (ligases). These were coupled with the polymerase chain reaction, a technology that allowed the precisely controlled multiplication of individual DNA strands. This not only facilitated analysis of the DNA being investigated, but it gave molecular biologists large quantities of specific DNA constructs that were then used for insertion into target organisms. The third technology discussed was the development of computers and communications systems in the 1990's that allowed for gene sequences to be shared among researchers and made available in remotely accessible computer databases.

        These three elements were the prerequisites for the large scale sequencing projects underway at that time, including the human genome sequencing effort. It is interesting to note that, while I went on at some length regarding gene function and activity, and the potentially immense power of genomic analysis, nowhere in the entire section do I use the word "proteins". One passage is telling: "If we could look at all the genes at the same time, we would know what's happening to all the genes at the same time - which are being upgraded or downgraded, which are being turned on or off, accelerated or decelerated." (10). In fact, this is a description of genomic/proteomic analysis, but as the word "proteomics" had not yet been coined in 1999, the passage only refers to analysis of gene expression. Scientific terms arise to describe new phenomena and new fields when old terms become unworkably clumsy.

        It is also interesting to note that this complete analysis of all genes and their expression was presented as a hypothetical technology. Taking a biochemical snapshot of the relative expression levels of the entire genome, estimated at that time to be at least 30,000 genes in many crop plants, and expected to be more than 80,000 genes in the higher vertebrates, was a daunting technical challenge. In fact, it was in the late 1990's that advances in DNA and protein technology were being turned into tools which would allow for just this kind of simultaneous analysis of multiple genes. This technoloy allows one to determine which genes are active, and to what degree, under various conditions. Since its introduction, it has served to accelerate understanding of the behavior of the entire proteome, and is what could be considered the fourth essential technology of the era of functional genomics. This technology went by an number of names before the common term "microarray" was adopted.

Microarrays - the Genome on a Chip

        Microarrays are wafers or chips on which have been spotted, in regular rows and columns, a number of tiny pieces of different binding agents. Some microarrays are designed to work with DNA, others with proteins, or with other cellular constituents. In the case of DNA microarrays, this agent is a piece of DNA that has the specific sequence that represents a particular known gene. Since their sequences are different, each spot binds very well to its complementary sequence and poorly to any other. To use the microarray, sets of cells being compared are grown under experimental conditions which cause different genes to be expressed. The mechanism of this expression is the transcription of DNA (the gene sequence) into messenger RNA (mRNA), which is then translated into an amino acid sequence (the protein gene product). The functional significance of this different expression is that mRNA will be present in the cell for genes that are active, but not present for genes that are inactive. The total mRNA is collected from the cells being tested; each strand of mRNA is used as a template to make a strand of DNA with a complementary sequence. This complementary DNA (cDNA) will bind to the pieces of DNA that are spotted on the microarray.

        A variety of technical means are available to determine when binding has taken place, such as changes in electrical conductivity, spectral absorbance or fluorescent color. In microarrays that use color-based separation, the cDNA from one set cells is bound to one fluorescent dye (e.g. green), and the cDNA from another set of cells is bound to another dye of a different color (e.g. red). The cDNA from all of the cells is combined and washed across the surface of the microarray. If a piece of cDNA with a given spot's complementary sequence is present in the mix, it binds; otherwise, the spot remains unbound. The spots are illuminated by a laser specific for the green dye, and all spots binding cDNA dyed green light up. The process is repeated with a red laser. The cDNA binding to each spot could have come from either of the types of cells being examined, or from both. A computer reads which binding sites light up under which lasers, and gives a readout of which spots light up under the green laser, which light up under the red laser and which light up under both. When a spot lights up, that means that a gene is active. If it is lit by one laser and not the other, that means that the gene is on under one set of conditions and off under the other. A spot lit by both lasers is active under both conditions; spots which are not lit under either represent genes that are inactive. The data is stored in the computer, and cross-referenced against a database of the genes that the microarry was designed to test.

        When microarray technology was first developed in the late 1990's, each wafer held around 40 spots, each with a different binding agent. Each wafer could, therefore, measure the expression of up to 40 genes simultaneously. This limitation meant that microarrays were designed to investigate only certain select groups of genes, customized to serve different fields of inquiry. Within a few years, however, the number of binding spots on the wafer, and therefore the investigative power of the microarray, increased in density tenfold, and then again by increasing orders of magnitude. In 2005, one commercially available DNA microarray system designed for use with the human genome will show, on a single chip, the simultaneous activity of 29,098 genes found in human cells (6). Similarly dense microarrays based on completed or partially completed genomes of other species (e.g. rat, fruit fly, rice, corn, etc.) are also available. These microarrays have been referred to as a "genome on a chip", a description that is now less marketing hype than simple statement of fact. With these microarrays, a scientist can know what every single gene in the cell is doing at any given time, and know how the cells change their response under changing conditions. With essentially the entire complement of genes available for analysis, the same microarray can be used for almost every field of inquiry, greatly facilitating standardization and sharing of data among researchers.

        The use of microarrays in biology is becoming as commonplace as the use of computers. Their introduction was a revolution; their continued use is an evolution. This approach is one of the underpinnings of the era of functional genomics, and informs investigations of human disease progression, clinical and veterinary drug discovery and testing, identification and development of new agrochemicals as well as screening and breeding of livestock and crops. Food production and distribution systems increasingly rely on microarrays, genomic and proteomic analysis and other types of biotechnology to improve efficiency and profitability.

The Biotechnology of Food

        In 2000, the Institute of Food Technologists published an Expert Panel Report on biotechnology and foods (7). This report, written by 28 top researchers, treats biotechnology in a fairly focused way, addressing issues related to the use of gene insertions in livestock and crops plants used for food, feed and fiber. The safety and wholesomeness of modified foods comes under consideration, as do issues related to their regulation, labeling, and marketing. As the authors of this report acknowledge, "one of the difficulties in discussing the benefits and concerns that attend any technology is consideration of the rapid and extensive advances". In the field of biotechnology, documents become obsolete almost before they are published, but with that caveat in mind, it is clear that many of the positions presented in this five year old report stand the test of time.

        The authors note that the use of recombinant DNA (rDNA), i.e. the combining of genetic material from disparate sources, is an extension of the genetic modifications that began with selective breeding at the dawn of agriculture some 10,000 years ago. The modern tools available increase the speed and precision with which traits can be identified in one organism and incorporated into another. Genetically modified crops can be bred to better withstand environmental stresses, better utilize soil fertility, resist pests and diseases, and give higher yields of better tasting, more nutritious foods which do not spoil in storage. Many of these observations were also made at the 1999 ITEST workshop (10), with the same cautions about confabulating hypothetical applications with practical ones.

        From a food processing standpoint, many enzymes and biochemicals with important food industrial applications have been traditionally derived from scarce or inconsistent sources, such as the enzyme chymosin. Chymosin is used by the dairy industry as a means of clotting milk, the first step in cheese production. Historically, this enzyme was extracted from rennet, taken from the stomach lining of young calves. In the 1990's, a number of companies competed to engineer bacteria, yeast or molds to produce the enzyme. Today, most cheese manufacturing uses this bioengineered product, not only because it is a fraction of the cost, but because its purity (>95% chymosin) is far greater than that of traditional rennet (~2% chymosin) (7). A host of other examples exist where industrially useful enzymes, once difficult to obtain in bulk, are now produced in commercial quantities by bioengineered bacteria or fungi. These are used to process corn into high fructose corn syrup, speed beer and wine fermentation, improve the aroma of jams and jellies, and many other applications. These enzymes, when used in food manufacture, must be approved as a GRAS (i.e. Generally Regarded As Safe) food additive by the US Food and Drug Administration (FDA).

        As with food-grade enzymes of unprecedented purity, when a new rDNA organism is created, it must pass regulatory approval before it can be used for food or feed. The US FDA has the authority over the introduction, labeling and use of new foods and animal feeds, including those created via biotechnological means. Within this context, then, "new" foods are treated the same as "old" foods, in that they are subjected to the same or greater levels of scrutiny. Is the food safe to consume or feed to livestock? Does it contain harmful chemicals? Is there an increased toxicological risk? Are new allergens present which might lead to adverse reactions? The FDA's authority extends over all food products, with the exception of meat and poultry, which are regulated by the US Department of Agriculture (USDA).

        Currently, few if any genetically modified animals have been proposed for approval by either the USDA or the FDA. In contrast, a number of crop plants have been approved and have become mainstays of US agriculture. Some of the best known examples are varieties of corn, sugar beets, cotton and canola that resist the herbicide glyphosate, and varieties of potato, tomato, corn, and soybean that produce a toxin derived from a soil bacterium. More than half of all soybeans grown in the US are genetically modified. Of the approximately 200 million acres of arable land planted with genetically modified crops worldwide, approximately 120 million is in the US, and almost 75% of all processed foods consumed in the US contains one or more genetically modified ingredient (3,9) The FDA's regulatory approval process for GMOs should be seen as one contributing factor for the extensive adoption of these crops in the US. Appearing on the horizon, however, is a change in the way foods are produced and marketed, which is challenging this approval process in ways never before explored.

Nutrigenomics: How Foods Change Our Gene Expression

        The production and distribution of wholesome food is the cornerstone of every society on Earth. The research of the last century brought a clearer understanding of the role of vitamins, minerals, lipids, fiber and other food components in a healthy diet. In many societies, even when total caloric intake was adequate, diseases resulting from deficiencies in necessary minor components were common. Today, diseases of malnutrition such as goiter, rickets and pellagra are essentially unknown in the Western world today. This is due in part to more balanced diets, but also to the degree to which so many of our foods are fortified with vitamins and minerals. Although in many parts of the world, starvation and malnutrition in their most basic forms are an ongoing concern, in industrialized nations the focus is shifting to the relatively subtle impact of minor food components on long-term health and well-being.

        Foods contain a variety of bioactive compounds which exert an influence on our metabolism. These compounds represent less than 1% of the food we eat, but may play a significant role in our propensity to develop chronic illnesses, such as heart disease, cancer, and arthritis. The mechanisms by which these minor constituents act is through interactions with transcription factors within our cells. When a compound binds to the promoter region of a gene, it may result in either increased or decreased production of that gene's product, which in turn may serve to regulate the functioning of one or more additional gene systems. The presence of food constituents can therefore lead to a cascade of interaction involving dozens, if not hundreds of genes.

        What does this mean for the everyday experience of choosing which foods to eat? It may very well be that consuming a diet rich in a beneficial bioactive compound may serve to protect against illness. Researchers have already identified an extensive list of receptors within our cell nuclei that are capable of binding different forms of fatty acids (4). Various genes may be turned on or off, depending on the level and type of fatty acids to which the cell is exposed. An example is the consumption of omega-3 fatty acids, found in certain types of fish and soy oils. Clinical research has shown that a diet rich in omega-3 fatty acids may be of benefit to people at risk of cardiovascular disease. The mechanism for this benefit, however, is not clearly understood. Using the microarray technology discussed earlier, scientists have measured the simultaneous expression of more than 12,000 genes in mice fed with diets rich in omega-6 or omega-3 fatty acids. More than 300 genes showed differential expression as a result of the diet (4).

        While results from animal studies are informative, as of this writing, human studies are still lacking. Where work has been performed measuring the expression of the human genome, these studies typically involve exposing a culture of human cells to the bioactive compound in a relatively purified form, and performing a microarray analysis of the expressed mRNA profile. Studies of this type have already shown some response of some food-associated compounds. Theaflavins, found in black tea, have been shown to reduce the expression of the COX-2 gene, which is one of the genes responsible for inflammation of tumors and arthritic tissues. Similar suppression of COX-2 has been shown by resveratrol (found in grapes), catechins (from green tea) and vitamin E (4)

        These studies are only a first step in nutrigenomic research. For foods to have pharmacological effects, the bioactive compounds must first be present in pharmacologically meaningful levels in the foods, and within a context of food chemistry that makes them biologically available to be absorbed. Then, they must not only survive the digestive processes of the stomach and gut, but must be able to be translocated throughout the body so as to reach the target tissue. Furthermore, they must arrive at the target site in an effective concentration, and in a chemical form which preserves the compound's original efficacy and potency. All of this takes place within the context of the human body. The goal is to influence the behavior of a tiny subset of the many hundreds of distinct cell types, without disturbing any of the others. Each of these cell types are expressing their own suite of genes, within their own distinct expressed proteome. To complicate matters further, each is operating within an ever changing flux of hundreds of thousands of different chemical compounds.

        Clearly, before one can draw conclusions regarding, for example, the anticancer benefits of drinking black tea, in vitro results must be validated with clinical trials to establish real-world efficacy. The advantage that genomics/proteomics technology provides is that, as a preparatory step, it gives scientists an indication of what to look for in the clinical trials with regard to the functioning of specific genes. In a feedback-type interactive system, the clinical trials can then inform the next round of in vitro tests and genomic/proteomic screening. The increased precision of knowing with greater accuracy exactly how these foods are influencing our gene expression will greatly improve our ability to draw meaningful conclusions and make efficacious recommendations.

Next Up: How Our Gene Expression Changes Our Food

        Nutrigenomics investigates how our foods change our gene expression. Once the mechanism is elucidated for how Compound X reduces the likelihood of Disease Y, it will surely not be long before Compound X starts showing up everywhere. Nutrigenomics is a field still in its infancy, but dietary guidelines and recommendations based on conventional clinical trials are being used to guide a new generation of bioengineered foods. Many of the latest "nutraceuticals" have, of course, been present all along in the foods we eat. However, once a clinical benefit has been shown, it frequently becomes a marketing tool. For example, lycopene is a fat soluble antioxidant compound found in tomatoes, watermelon, grapefruit and some other characteristically red fruits. In recent years, diets rich in lycopene and other antioxidants have been suggested to help reduce rates of various types of cancer. The efficacy of lycopene is best seen when it is ingested with foods, rather than as a stand alone dietary supplement. In response to these findings, products ranging from spaghetti sauce to catsup proudly proclaim, "Now with Lycopene!"

        If a food is truly being consumed to achieve a pharmacological benefit, then as a first step, the active compound must be present in pharmacologically meaningful concentrations. One of the ways to ensure that this concentration is present is to create foods that produce more of the compound, or by engineering away extraneous compounds which may interfere with the bioavailability of the compound of interest. Biotechnology tools that enhance the analysis of existing genetic structures are firmly entrenched in modern plant breeding; new varieties that express a desired trait, such as tomatoes with enhanced lycopene content, can be produced far more quickly than in the past, even without the introduction of rDNA. It is relatively simple to go one step further and insert a few extra copies of the specific genes responsible for production of the desired nutraceutical compound, to perhaps double or triple the levels in the fruit or vegetable.

        When an idea becomes entrenched in the marketplace, products evolve to take advantage of it. Medical authorities such as the National Institute of Health have recently begun promoting a catchy slogan to improve dietary choices, "5 A Day The Color Way". The goal of the promotion is the encourage people to eat fruits and vegetables that are colorful (red, orange, yellow, purple, etc.). It is based on research that shows the potential benefits of increasing the intake of phytochemicals, the different compounds that give these food their colors. When science is used for marketing (i.e. "red fruits = lycopene = healthy heart"), comparisons will inevitably follow to determine the relative merits of one functional food vs. another, such as how much lycopene is actually in various kinds of catsup. For example, one recent study showed that catsups which were darker red had more lycopene (8). It's not hard to speculate on how this information could be used in a marketing campaign: "More lycopene than the others... you can See The Difference!" "Eat more lycopene to help your heart - Better Red Than Dead!".

        When an attribute is seen as especially healthy and desirable, what about products which can't take advantage of the buzz? Many red fruits, such as cherries, strawberries, and raspberries, owe their color, not to lycopene, but to anthocyanins, a completely different class of compounds. Anthocyanins have their own purported nutraceutical properties, but they are still a matter of ongoing research. The same limitation could be presented for any functional food product which has one good attribute, but not all good attributes. To get the best of both worlds, fruits and vegetables which are rich in one class of compounds may be bioengineered to produce both, thereby going from being merely functional food to being multifunctional.

        The concept of using foods to improve health is certainly not a modern invention. However, the way in which the concept is evolving, particularly with regard to the genetic modification of foods, suggests something more complex than mere healthy eating. Functional foods and nutraceuticals are offered as a means to adjust the behavior of specific genes within our cells. Genomics and proteomics analysis will allow us to determine the exact extent to which specific diets achieve that goal. Extending the concept would have us altering our food intake and ultimately altering the foods themselves in order to enhance their effect on gene expression. As the functional foods movement gains ground, food may increasingly be taken for pharmacological rather than nutritional benefit. In contemplating a society where food is not really food, one is reminded of Lewis Carroll, if not George Orwell.

        Herbal supplements are widely popular, because they are seen as being a "natural" means of changing how our bodies function. This entire class of products is not well regulated. In contrast, direct means of changing our gene expression are regulated, controlled and regarded with caution by most reputable scientific bodies. One method, gene therapy, involves providing a patient with functional copies of a gene which he or she lacks. The hope is that the functional gene will become incorporated into the patient's cells and begin to provide whatever cell component was lacking, thereby curing the patient's disease. In numerous trials, dealing with a variety of diseases, this approach has resulted in some benefit, much inconclusive data and several deaths. The proteomic regulation of genes within the human body is still too poorly understood to allow for this kind of alteration of gene function. How is it, then, that indirect manipulation of our proteome, through supplements or functional foods, seems more acceptable?

        The answer, I believe, is that discussions of exactly how these nutraceuticals promote good health are not what drives acceptance. In the industrialized world, consumers have become accustomed to seeing our foods modified, amended and fortified so as to improve their healthfulness. Vitamins and minerals are added to thousands of food products, from cereals and bread to milk, juices and even bottled water. It seems a natural extension then, when a new compound is identified as being healthy, to add it to our processed foods. However, minimally processed foods, such as fresh fruits and vegetables, have had to stand on their own merits, offering as health benefit only that combination of phytochemicals endowed them by their breeder. Adding to the list of compounds they present will serve to make these foods even more effective agents of good health. Even when health claims have not been subjected to scientific validation of their mechanism or demonstration of actual benefit, these products become widely adopted. Consumers are not so much concerned with the mechanism of the benefit (alteration of our gene expression) as in the benefit itself (lowered risk of disease).

Benefits and Mechanisms

        This focus on selling the benefit, rather than the mechanism, is what will drive the increasing adoption and application of genetic manipulation in a variety of life sciences. Biotechnology is a firm fixture in areas of our society on the periphery of ethical consideration, dealing with plants and plant products. Bioengineered species are used extensively in the production of crop plants for food, feed and fiber, as well as in a number of other applications. Horticulturalists have been unsuccessful in hundreds of years of traditional breeding attempts to create a truly blue rose. This feat was recently accomplished using a complex package of genetic insertions (1). First, the genetic pathways responsible for the synthesis of the rose's native palette range of reds, yellows and oranges (the DFR gene) had to be turned off. This was accomplished by inserting a synthetic gene which produced an mRNA product which was the mirror image of the mRNA produced by the DFR gene. This "anti-sense" mRNA bound with the DFR mRNA, preventing it from being translated into a protein, effectively silencing the DFR gene. With the native red pigment pathway blocked, a pathway to create a precursor for the blue pigment had to be introduced. A gene isolated from pansies, called delphinidin, was inserted to support the new pigmentation. Finally, to actually make the blue pigment, a different form of the DFR gene obtained from irises and inserted into the rose. This triplet package of genes results in a truly blue rose, a prize worth millions of dollars in horticultural circles.

        What was learned from producing this rose has broader applications (1). The pigment precursors in conventional roses are modified to eventually result in a red palette; the bioengineered roses silence the pathway to produce the native precursors, add a pathway to make novel precursors and add a new modification enzyme. In broad terms, this technology could potentially be be used to change the mixture of precursors and final cellular components in a range of plants. Research is already underway to alter the lipid profile of oilseed crops such as cotton, flax and canola so as to produce omega-3 fatty acids and other valuable oils. Production of plant-derived flavorings and essential oils could also be enhanced by this type of modification.

        Another example is the modification of existing crop plants to enhance their utility as biofuels. "Biotechnological Enhancement of Energy Crops" is the title of an ongoing research project at the USDA (2). This multiyear project is specifically geared toward using genomic and proteomic tools to determine the genetic controls on how plants capture and store energy, and developing ways to alter these controls so as to improve their potential for use as a supplement or replacement for fossil fuels. This might involve changing the plant's chemical composition or anatomical architecture to improve sunlight capture efficiency. Additional research is investigating the use of new enzymes, reagents and biocatalysts to improve the efficiency with which biofuel feedstocks are processed industrially. Similarly, processes to convert byproducts with low economic value into more valuable commodities are being tested. By increasing the percentage of usable ethanol, biodiesel or other fuels from a given amount of corn, soy, switchgrass or other feedstock, or by otherwise increasing the value of the derived products and byproducts, the economics of biofuel production can be improved.

        Even when limiting the discussion to plant bioengineering, many other examples are available as to how biotechnology is being applied. Increased pest resistance, spoilage suppression, herbicide tolerance, plant-based vaccines... the list is long and grows longer every year. These advances are not universally accepted. A vocal minority seeks assurances that their food and feed products are "GMO free". Still, as the percentages of GMO crops increases steadily, it will eventually be the non-GMO crops that are specialty items. It seems likely that, as the benefits of bioengineering become ubiquitous in plant sciences, animal sciences must eventually seek to obtain the same benefits.

        While biotechnology is already used extensively in analyzing the genetics of animals to support breeding programs, direct intervention by genetic modification is still limited. Recent studies have shown that the milk and meat derived from cloned cattle are identical to that obtained from conventional cattle (12). It is a measure of the controversial nature of the technology that this result was met with a renewed firestorm of debate. The cloning of livestock prompts concerns regarding the potential dangers of monocultures, the economic position of small family farmers vs. large corporate farms, and the morality of adopting such a high-tech reproductive tool that has, at best, only a 1-3% rate of embryos resulting in successful births. On the other side of the debate, the potential increases in milk and meat production, the improved quality of wool and leathers, and the ability to create sturdier, more disease resistant and robust livestock would seem to be to society's benefit.

        The distinction in how biotechnology-derived plants and animals are viewed is not purely academic. The FDA is currently reviewing the data on cloned animals, and is expected shortly to issue a ruling on the safety of food derived from them. The protocol which the FDA follows in its regulation of genetically modified crops is relatively permissive, and will act to pull a plant product from the marketplace if a problem is identified in its use. If a more conservative stance is adopted, the FDA may require increased toxicological testing prior to marketing, additional labeling requirements, separate packaging and shipping of bioengineered animal products from traditional products, or other actions. However promising the science may be, too many political or economic hurdles may prevent the adoption of these technologies.

        That statement also summarizes, in brief, application of biotechnology to humans. Screening of existing genomic expression to detect genetic diseases, or changes following some pharmaceutical or nutraceutical treatment are tolerable; direct interventions to change our genomic expression may not ever be. These technologies as they might be applied to humans are still, at this point, firmly in the realm of the hypothetical. As the potential bioengineering of humans looms larger on the horizon, however, the discussion has led to state and federal regulations to provide for proactive ethical guidelines and limits on research. It has also led some states to provide extra funds to support this research, in hopes of being in the vanguard when the next wave of biotechnology breakthroughs becomes the industries of tomorrow.

        The cooperation and competition of public and private researchers laid the foundations for the science of genomics and proteomics. The societal pressures of interested parties on all sides of the debate is shaping the legislative framework for how this science will be applied to animals and humans in the coming century. The ever-advancing technical state of the art means that the transition from the hypothetical to profitable with extreme rapidity. It is not possible to come to a final decision regarding how we should deal with these advances, because the technical situation is in a state of constant flux. Guiding the development and adoption of biotechnology in a way that benefits people as individuals and society as a whole will require attentive, thoughtful and, most importantly, continuous reevaluation.

References

1. Anon. 2005. "Plant gene replacement results in the world's only blue rose". PhysOrg.com www.physorg.com/news3581.html (Accessed Aug 24, 2005)
2. Biotechnological Enhancement of Energy Crops. USDA-ARS Project Number: 5325-21000-013-00. Project Description: www.ars.usda.gov/research/projects/projects.htm?ACCN_NO=408875 (Accessed Aug 24, 2005)
3. Brubaker, H. 2005. Philadelphia Inquirer. "Spurning Biotech" p. C1, June 17, 2005
4. Hirch, J.B. and D. Evans. 2005. Beyond nutrition: the impact of food on genes. FoodTechnology 07.05:24-33
5. Human Genome Project Information (HGPI). 2005. www.ornl.gov/sci/techresources/Human_Genome/project/info.shtml (Accessed Aug 19, 2005)
6. Human Genome Survey Microarray v2.0. (HGSM) 2005. Applied Biosystems, Foster City, CA www.appliedbiosystems.com (Accessed Aug 22, 2005)
7. Institute of Food Technologists (IFT). 2000. "IFT Expert Report on Biotechnology and Foods" members.ift.org/IFT/Research/IFTExpertReports/biotechfoods_report.htm (Accessed Aug 23, 2005)
8. Ishida, B.K., Chapman, M.H. 2004. A Comparison Of Carotenoid Content And Total Antioxidant Activity In Catsup From Several Commercial Sources In The United States. Journal Of Agricultural And Food Chemistry. Vol. 52. P. 8017-8020.
9. Johnson, L.A. 2005. Philadelphia Inquirer. "Genetically modified foods more common" p. A8, March 25, 2005
10. Niemira, B.A. 2000. Cellular pesticide production in genetically modified crop plants: biochemistry, economics and intellectual property. pp. 19-48 in Brungs, R. and S.M Postiglione (eds.) The Genome - Plant, Animal, Human. ITEST Faith/Science Press, St. Louis, MO.
11. Roberts, L., Davenport, R.J., Pennisi, E., Marshall, E. 2001. A History of the Human Genome Project. Science 291: 1195
12. Tian, XC, Kubota C, Sakashita K, Izaike Y, Okano R, Tabara N, Curchoe C, Jacob L, Zhang Y, Smith S, Bormann C, Andrew S, Yang X. 2005. Meat and Milk Compositions of Bovine Clones Compared with Matched Controls. Proc Natl Acad Sci USA 102: 6261-6266.

Copyright 2006, Brendan A. Niemira