Everyone wants the human genome to be a turnkey solution to human health. All diseases have their basis in genes and proteins, after all--even if they're an invader's genes and proteins--so it doesn't seem farfetched to believe that the catalog of human genes could yield the long-sought after "magic bullet."
Monoclonal antibodies once flirted with this tag, but immune responses and the complexity of biological systems disappointed. Antisense RNA, which binds to messenger RNA (mRNA) and prevents it entering the cell's transcription machinery, also had its time in the spotlight. But antisense, too, fell victim to pitfalls, including single-stranded RNA's notorious instability and a tendency to activate a nonspecific interferon response toxic to cells.
In 2002, Science announced the official arrival of the latest contender: RNA interference (RNAi). It bears superficial resemblance to antisense, which despite shortcomings has advanced several drug candidates to the clinic. But RNAi works by an entirely separate mechanism, a natural cellular defense against viral infection. Long double-stranded RNA is a sign of viral infection, and cells respond by recruiting an enzyme called "dicer" to chew it up into smaller pieces. These bind to a protein complex known as the RNA-induced silence complex (RISC), which removes the noncomplementary strand and uses the remaining one as a template to guide the digestion of target mRNA. One of the strengths of RNAi is that the RISC process is catalytic, so that interfering RNA can have long-lasting effects because RISC reuses it.
Initially, RNAi seemed to work only in nonmammalian organisms such as C. elegans. But in May 2001, researchers at the Max Planck Institute for Biophysical Chemistry published a paper in Nature (S. M. Elbashir, J. Harborth, W. Lendeckel, et al., "Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells," Nature 411, 494) demonstrating that short sequences (known as small interfering RNAs, or siRNAs) effectively shut down protein production. In fact, siRNAs are small enough that dicer ignores them.
The timing of that announcement couldn't have been better. The February 2001 publication of the first draft of the human genome had researchers scratching their heads over the best way to tease apart the functions of the genome's estimated 30,000 genes. Doing so requires knocking out a gene and observing the effect on phenotype, but the few methods available to do that, such as antisense, ribozymes, or aptamers, were painstaking or imprecise, or both. Enter RNAi. "It's part of that next wave of postgenomics technologies ... it bridges genomics and proteomics," says Robin Rothrock, director of market research at BioInformatics LLC in Arlington, Virginia, and author of a recent RNAi market report.
The biotechnology industry has wasted no time in adopting RNAi for target identification. "It allows us to do large-scale gene knockdown experiments. That type of technique was not available to us previously," says Stephen Fesik, divisional vice president of cancer research at Abbott Laboratories in Abbott Park, Illinois. So-called rational drug design is dependent on some degree of certainty that a receptor or enzyme is a reasonable target for a drug. It's best to find that out early in development so that a company can focus its resources on the candidates most likely to produce a payoff. Although it has yet to be conclusively demonstrated to function in an animal model, RNAi makes it easy to verify--in cellular assays, at least--whether or not a given protein plays an essential role in a specific disease pathway.
A nascent industry has appeared to provide those services. Companies like Ambion Inc. in Austin, Texas, and Proligo LLC in Boulder, Colorado, are broadening their traditional base as reagent suppliers to include siRNAs and other reagents for RNAi research. Others are contracting with companies like Abbott and Merck to provide target identification, or building libraries of optimized siRNA reagents. In June, Lafayette, Colorado-based Dharmacon Inc. signed on with Exelixis Inc. in South San Francisco, California, to produce siRNA reagents that target 600 popular drug target candidates, including kinases. Dharmacon is currently delivering to Abbott an expanded library of 4000 siRNAs that target kinases, G-protein coupled receptors, and ion channels, says Fesik.
These research products represent one arm of RNAi. The other is drug development. The double-stranded RNA that accomplishes RNAi is more stable in vivo than the single-stranded RNA of antisense. It also is less toxic than antisense, although a couple of recent reports have suggested that RNAi may, like antisense, activate nonspecific immune responses.
Where the Jobs Are
RNAi is far from perfect, and it is the warts that figure to attract the biggest investments--and the lion's share of new jobs-- in the coming years. When it comes to therapeutics, the biggest obstacle is "delivery, delivery, delivery," says Dmitry Samarsky, director of technology development at Sequitur Inc. in Natick, Massachusetts.
Although more stable than antisense, the double-stranded RNAi molecules are nevertheless metabolized and excreted. But targeting the agents to cells should circumvent such problems, and researchers would like the effect to be as specific and prolonged as possible. Companies are taking various approaches to the problem--some are experimenting with various vectors, while others tinker with the structure of RNAi itself, modifying it in hopes of slipping under the immune system's radar screen.
It would also help to get a better handle on the complex RISC complex. "We don't know how many proteins are in there, or what each one does," says Nassim Usman, who is chief scientific officer and vice president of research and development at Sirna Therapeutics in Boulder, Colorado.
Although many of the jobs in the RNAi industry will closely resemble positions in other industries--biochemists, molecular biologists, and pharmacologists will be in demand--some specializations will be particularly sought after. Naturally, familiarity with nucleotide chemistry is a plus. "We need general nucleic acid chemists who know not only how to modify and make RNA, but also how to formulate and deliver nucleic acids, which is a somewhat obscure field. People don't really train you to do this," says Usman.
Not everyone agrees that there is an RNAi industry at all. Companies have sprouted up to take advantage of the new technology, but most of its industrial application comes from larger biotechnology and pharmaceutical companies who see it as an important tool in drug discovery.
Still, reagent suppliers have expanded their operations to produce additional reagents, and they will no doubt be a source of new employment as they continue to develop new products and expand their lines in answer to evolving research demands.
Other companies foresee the day when RNAi compounds will fulfill the clinical promise that remains theoretical. The first human studies are likely still at least 2 or 3 years away, says Christoph Westphal, who is a general partner at Polaris Venture Partners in Waltham, Massachusetts. Westphal was the founding CEO of Cambridge, Massachusetts-based Alnylam Pharmaceuticals, which is developing RNAi therapies, and Polaris holds a stake in the company.
Mergers and acquisitions are likely in store, as are more start-ups and IPOs. As soon as RNAi shows efficacy in animals, large biotechs and pharmaceutical companies will pour more money into RNAi companies in the form of collaborations and licenses, says Westphal.
Therapeutic applications are likely to hit easy targets first, says Nagesh Mahanthappa, director of corporate development at Alnylam. "The near-term advances will be opportunistic, such as taking advantage of some intrinstic ability of siRNAs to accumulate in specific cell types or tissues. Another [approach] is to make use of tissues or sites where one can administer siRNA without fear that broad distribution will result in degradation and loss of activity. Companies have talked about the eye, and there's precedent for injecting directly into eye," says Mahanthappa. Other possibile targets include the lungs and skin.
Patents may also become an important factor, according to Charles Chang, a strategic market report analyst at Front Line Strategic Consulting Inc. in San Mateo, California. Several important patents are currently being disputed, and a number of companies continue to work in the field, using methods that they may soon have to license. "When the patent disputes get settled, certain companies that don't have [rights to key technologies] will exit the market or be acquired by larger players that have rights to siRNA technology," says Chang.
But for those just looking for a job, the patent outlook is too murky to worry about. Rest assured that by all accounts, the future of RNAi is secure. How big will it be? That remains open to question.
Westphal thinks RNAi may vault to the top of biotechnology's key discoveries, joining recombinant DNA and monoclonal antibodies as a pillar of 21st century medicine. "This could be a third very important discovery for the basic understanding of molecular biology," he says.
But Westphal is admittedly biased. Abbott's Fesik is more reserved. "This is one of many technologies we're excited about" to assist in early stage drug discovery. And that's a critical stage. "If you are not working on a target that is validated, then everything you do downstream of that could be a waste of time."