PROTEOMICS
Can Celera Do It Again?
Robert F. Service
The upstart genomics firm is set to embark on an ambitious new effort in protein analysis, but it faces a huge challenge and some stiff competition from companies already in the field
J. Craig Venter and Michael Hunkapiller proved the skeptics wrong once. In 1998, the two teamed up in a bold plan to sequence the human genome by the end of 2001, a full 4 years ahead of the finish date then projected by the publicly funded Human Genome Project. Venter, head of Celera Genomics Corp. in Rockville, Maryland--the firm created to carry out this mission--was the brains behind a novel genome sequencing approach. Hunkapiller, head of the scientific equipment powerhouse PE Biosystems, provided brawn: a suite of the company's experimental high-speed DNA sequencers. Despite strong doubts among genome researchers that Celera could pull it off, the project appears to be just months away from completion.
Now the pair is preparing to push Celera beyond the genome to conquer the next frontier--the identification of proteins involved in human disease. To do so, Celera is launching a major effort in "proteomics," an effort to identify all the proteins expressed in an organism and then track their ebb and flow. The move, says Venter, is the next logical step in understanding the role of all the genes they've decoded. It is also a critical step in developing novel drugs and tailoring medical care to the genetic makeup of individuals. To pay for the new initiative, Celera raised $944 million in a stock offering earlier this month, much of which will be devoted to proteomics.
This time, however, Venter and Hunkapiller are tackling a much more complex problem, and they will be facing competition from companies ranging from small start-ups to big pharma that have already started proteome projects of their own. "I don't think they are more credible than anybody else," says Mattias Mann, chief scientific officer of Protana, a proteomics company in Odense, Denmark.
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KEY CORPORATE PLAYERS IN PROTEOMICS
Company Location Approach
Celera Rockville, MD Databases
Incyte Pharmaceuticals Palo Alto, CA Databases
GeneBio Geneva, Switzerland Databases
Proteome Inc. Beverly, MA Databases
PE Biosystems Framingham, MA Instrumentation
Ciphergen Biosystems Palo Alto, CA Protein arrays
Oxford GlycoSciences Oxford, U.K. 2D gel/MS*
Protana Odense, Denmark 2D gel/MS
Genomic Solutions Ann Arbor, MI 2D gel/MS
Large Scale Proteomics Corp. Rockville, MD 2D gel/MS
* 2D gel electrophoresis and mass spectrometry.
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But Venter, never one to understate his ambitions, boasts: "We're going to dominate in our own way. We're going to have the biggest facility and the biggest database." He concedes that the huge amount of work needed to understand proteins and their interactions inside cells guarantees that many academic labs and other companies will also be players in the field. But, he says, "we're building a Celera-scale proteomics facility" capable of identifying up to 1 million proteins a day.
Plans for the new facility are still coming together, Venter says, but it will likely consist of a fleet of up to 100 machines, including high-speed mass spectrometers for protein analysis, as well as additional protein separation devices. Celera also plans to boost the capacity of its $100 million supercomputer --which currently holds some 50 terabytes of genome data--by a factor of 10 to handle the expected torrent of protein data.
Venter will have Hunkapiller's help getting his operation up to speed. Last week, PE--the parent company of both Celera and PE Biosystems--announced that it will form a proteomics research center at its PE Biosystems site in Framingham, Massachusetts, to create new high-speed machines. As part of that initiative, PE officials plan to pursue two technologies--one developed by Denis Hochstrasser and colleagues at the University of Geneva in Switzerland, the other by Ruedi Aebersold at the University of Washington, Seattle--that aim to do for protein analysis what the high-speed gene sequencers did for genome work.
Outside observers say that with PE Biosystems' backing, Celera's move into proteomics is likely to be pivotal for the emerging field. "The genomics company stuck its flag in the arena of proteomics," says William Rich, president and CEO of Ciphergen, a proteomics company based in Palo Alto, California. "It sends a message to the protein people that the gene people are not going to sit around and wait" for proteomics companies to give them the information they're looking for.
In fact, the march of genomics companies into proteomics is well under way. Incyte Pharmaceuticals, one of Celera's chief genomics rivals, is 2 years into an extensive partnership with Oxford GlycoSciences (OGS), a proteomics company based in Oxford, England. OGS creates protein profiles of different tissues in both healthy and diseased states, and Incyte incorporates this information into a proteome database that it markets to pharmaceutical companies. Incyte signed up its first subscriber to its proteomics database last fall, giving it "a significant lead over other companies in developing proteomics databases," asserts Incyte CEO Roy Whitfield. And like Celera, the company is also flush with cash. Incyte recently raised $620 million on the stock market, much of which is intended to bolster their proteomics work, says Whitfield.
Other companies are pushing into proteomics as well. Virtually every major pharmaceutical company has a proteomics effort under way, says Hanno Langen, who directs proteomics research at Hoffmann-La Roche in Basel, Switzerland. Moreover, small proteomics firms, such as Genomic Solutions of Ann Arbor, Michigan, and Large Scale Proteomics of Rockville, Maryland, are gearing up for initial public stock offerings to raise money for expanded research.
That makes Celera a late entry into the field, and it has some catching up to do. But it is betting nearly $1 billion that it can close the gap.
The next step
This move toward understanding proteins has emerged from the increasing recognition among genomics and pharmaceutical researchers that identifying DNA, or even messenger RNA (mRNA)--the nucleotide messengers that signal cells to produce a particular protein--is not enough. Neither DNA nor mRNA can identify how much protein is produced inside a cell or what it does once created. Although researchers initially hoped that the presence of a large amount of a particular mRNA meant that copious quantities of the corresponding protein were being produced, "there is significant evidence that there is not necessarily a correlation between mRNA levels and protein levels," says Philip Andrews, a proteomics researcher at the University of Michigan, Ann Arbor. Other factors complicate the picture as well, he adds. For instance, chemical modifications such as phosphorylation play a key role in controlling protein activity; these modifications cannot be detected by screening nucleotides. "The genome tells you what could theoretically happen" inside the cell, explains Raj Parekh, the chief scientific officer at OGS. "Messenger RNA tells you what might happen, and the proteome tells you what is happening."
Figuring out what's happening at the protein level won't be easy even for Celera. "Proteomics is a much more difficult problem than genomics," says Andrews. Whereas the human genome remains largely unchanged among individuals, he explains, the expression of proteins varies widely. Protein expression changes dramatically from one tissue to another and even within single tissues over time as a person ages. What's more, thousands of chemical modifications occur after proteins are created that alter their enzymatic activity, binding ability, how long they remain active, and so on. Although there may be only some 100,000 human genes, the myriad of modifications may give rise to 10 million to 20 million chemically distinct proteins in a cell, says Andrews.
This complexity, Andrews and others say, makes it almost meaningless to consider a human proteome project--akin to the human genome project--to identify all proteins in every tissue. The best researchers can do is try to focus on changes in key proteins, such as those involved in disease and development. For that reason, skeptics argue that even with Celera's deep pockets it will not be able to sweep aside the competition. "I think it will be very hard for any company to be the dominant proteomics company, much more so than in genomics," says Mann of Protana. Venter agrees--in principle. But he adds that few competitors will be able to match Celera's industrial approach. "We'll be working through every tissue, organ, and cell," he says.
Brute force
In the current proteomics rush, most companies are taking more or less the same brute-force approach to determining which proteins are present in various tissues, a technique called two-dimensional (2D) gel electrophoresis. Researchers start with a protein extract from a tissue of interest and then add it to a sheet of polymer Jell-O. By applying electric fields across the length and width of the sheet, they separate proteins by their electric charge and size. The result is a series of up to several thousand spots, each containing one or more types of proteins.
Once segregated into separate spots and stained, the proteins are typically cut from the gel one by one, chopped into fragments with an enzyme called trypsin, and dried. Then they are fed into a mass spectrometer that weighs each fragment, forming what amounts to a mass fingerprint of the protein's fragments. From that fingerprint, researchers can work out the likely combination of amino acids comprising it and then compare that to a genomics database to identify the corresponding DNA sequence. With the DNA sequence in hand, they can get a clearer identification of the protein. Researchers can then monitor changes in the expression of that protein to see whether it correlates with a disease state or perhaps a drug response.
All of this takes time. Today's top-of-the-line 2D gel operations, complete with robots to cut apart the gels and computers to analyze the information, can study a couple of thousand proteins a day. As a result, it can still take months to figure out which proteins change their expression in one set of tissues.
It's here that Venter and Hunkapiller hope to make a difference. PE Biosystems produces mass spectrometers, among other things. And according to Hunkapiller, mass spectrometers now in the prototype stage have the potential to go "orders of magnitude" faster than the current variety, making it possible to analyze hundreds of thousands of proteins per day. That scale of improvement in gene-sequencing machines enabled Celera and others "to ask whole new questions," says Hunkapiller. "If we can do the same thing in the protein area, it should have the same effect." By analyzing a large proportion of proteins in cells instead of just a select few, "you will see the things going on that maybe weren't so obvious," predicts Hunkapiller.
But this newfound speed will create bottlenecks upstream and require a faster front-end procedure to separate proteins and feed them into the high-speed mass spectrometer. This is where PE is looking to Geneva's Hochstrasser for help. Last fall, Hochstrasser and his colleagues published work on a new laser-based "molecular scanner" that automates the process of moving separated proteins from the gels to the mass spectrometers. The system applies an electric field perpendicular to the gel sheet; this field draws the proteins through two membranes. The first membrane is studded with trypsin, which digests all the proteins in a gel simultaneously as they move past. The fragments are then trapped by a second membrane, which is fed to the mass spectrometer. Finally, a laser marches along the membrane firing a steady stream of pulses in micrometer-sized steps, each of which blasts protein fragments into the mass spectrometer for fingerprinting. Here, too, the speed should reach tens of thousands of proteins per day, says Hochstrasser.
Working in tandem, the molecular scanner and a high-speed mass spectrometer could be a powerful combination, says Andrews. "The idea is spectacular. But whether it will perform as it needs to remains to be seen," he cautions. OGS's Parekh, for one, is skeptical. A big problem, he says, is how to quantify the amount of protein in each spot on the gel. This is necessary for identifying which proteins change in a stable way with disease--itself a first step to identifying molecular markers of disease and potential drug targets.
The time-consuming approach of gel cutting does allow such quantification. But that's typically not the case when the spots are transferred, or blotted, to another membrane, says Parekh. "Some proteins don't blot. Others are lost in the process. So as soon as you blot, you lose the quantity information," he says.
PE is betting that Aebersold's technology will help. In the October 1999 issue of Nature Biotechnology, Aebersold and his University of Washington colleagues reported a new approach that uses stable isotopes to quantify numerous proteins in cell extracts with mass spectrometry. The advance is "outstanding," says Hochstrasser, because it allows mass spectrometers to pin down protein levels from large numbers of proteins at once--a difficult proposition with today's technology. At this stage, it is not clear whether the new technique will work with a molecular scanner and a high-speed mass spectrometer. To find out, Aebersold is already working on joint research projects with PE scientists.
Even if it takes a while to get the mass spectrometers up to top speed, Celera can still make considerable progress, says Venter, as the company will be using other proteomics tools as well. A key approach, he says, will be to create antibodies to all proteins. These antibodies can then be used to fish out of a sample both targeted proteins and those they interact with. That, Venter says, will help Celera build up a database of how proteins interact with each other in complex biochemical pathways--information that is likely to be valuable to drug companies aiming to intervene in those pathways at specific points.
Incyte's Whitfield says he is not fazed by Celera's entry into the field. Even if PE and Celera manage to pull all these pieces together and launch a high-speed proteomics effort, other companies will also be developing their own high-speed approaches, he says: "We all understand that faster, cheaper, better is the way to go." With Celera preparing to enter the field, Whitfield adds, "I'm sure there is going to be great competition."
Volume 287, Number 5461 Issue of 24 Mar 2000, pp. 2136 - 2138
¸2000 by The American Association for the Advancement of Science.
Other great stories on celera:
highwire.stanford.edu
like this one:
BIOTECHNOLOGY:
How a Bland Statement Sent Stocks Sprawling
Eliot Marshall
Muddled news reports and a volatile stock market turned a presidential statement on genome data last week into a disaster for many biotech companies. Stocks of genetic research companies, after shooting upward early this year, plummeted on 14 March when President Bill Clinton and British Prime Minister Tony Blair issued a bland statement urging all labs to provide "unencumbered access" to raw DNA sequence information (Science, 17 March, p. 1903). Almost immediately, biotech stocks, which were already headed downward, went into a nose dive; some companies lost as much as 20% of their value on paper in a few hours. Within 48 hours many began to stabilize, but remained well below their peak a week later. Industry analysts had trouble interpreting these market gyrations. One biotech expert suggested a simple explanation: Stock buyers "don't understand what they're investing in," he said, and they can be easily spooked.
The spark that ignited the panic may have come during an informal briefing given by Clinton's press secretary Joe Lockhart on the morning of 14 March. As The Wall Street Journal reported the next day, Lockhart told a "gaggle" of regulars who cover the president that Clinton and Blair intended to issue a statement in the afternoon about a plan to restrict the patenting of human genes. If this is what Lockhart said--his remarks were off the record--it was not correct. Francis Collins, director of the U.S. National Human Genome Research Institute, says the statement was never meant to describe a new policy. The wording--which had been debated and revised "in many iterations ... over many months," Collins says--simply affirmed support for a 1996 research policy that calls for the immediate release of raw sequence data. Indeed, the Clinton-Blair statement specifically endorsed the patenting of "new gene-based health care products." But this clear message became tangled in stories of the rivalry between publicly and privately funded genome scientists over who should control human genome data (Science, 10 March, p. 1723). The upshot: Early news reports were confused.
At 9 a.m., CBS Radio News broadcast that the United States and Britain were aiming to "ban patents on individual genes." The Associated Press reported that there was a plan to restrict gene patents, but later said that Britain and the United States would begin to "openly share data" on the human genome. (They already do.) The stories became clearer later in the day. Even so, Chuck Ludlam, vice president of the Biotechnology Industry Organization in Washington, D.C., who saw the Clinton-Blair statement as "positive news" for industry, says he found it "unbelievable how wrong the reports were all day."
White House spokesperson Jake Siewert later told Science that "we completely dispute" the Journal's account of what caused the muddle. Lockhart, he says, told reporters that the Clinton-Blair announcement "had to do with public access to raw genomic data." But there was "confusion" during the "back and forth" between Lockhart and the reporters, Siewert concedes. "I don't think Joe got it perfectly right. ... And some reporters didn't get it perfectly right."
During the confused morning, stocks of companies that are creating private genetic databases--such as Celera Genomics of Rockville, Maryland, and Incyte Pharmaceuticals of Palo Alto, California--began to tumble. Other genome-related stocks began to slide, too. Soon the entire biotech sector slumped, as did the Nasdaq stock exchange index, which tracks high-tech firms. The Nasdaq index bounced back within 48 hours, but dropped again later, as investors remained wary of genomics and biotech companies. A week later, Celera and Incyte stocks, for example, were still 60% below their peak immediately before the statement. Predicts industry analyst Sergio Traversa of Mehta Partners in New York City, "Investors will remain a little bit more careful now," having been stung so badly.
- whatever is the proper disclaimer, I claim it. I think that if these magazines want to start saying that research should be made available for free, such as hgs, then they can't really start complaining about copyright violations, not that I think I am making one now. |
