Jan. 6, 1998 BERKELEY, CA. -- Carolyn Bertozzi and her colleagues at the Ernest Orlando Lawrence Berkeley National Laboratory have found a way to use natural biological processes to plant artificial markers on the surfaces of living cells.
With these markers, cell surfaces can be engineered to control cell adhesion to synthetic organic polymers, metals, ceramics, and other materials used in the walls of bioreactors, and in biomedical implants such as pacemakers and artificial organs. In the future, living cells attached to electronic devices may warn of dangerous chemical or biological toxins in the environment. Already Bertozzi's group has used this cell-surface engineering to turn cancer cells into bright targets for diagnostic probes and cell-killing toxins.
"Our primary goal is to take control of the cell surface," says Bertozzi, who is a member of Berkeley Lab's Biomolecular Materials Program in the Materials Sciences Division, as well as an assistant professor of chemistry at the University of California at Berkeley. "We have begun to understand the bio-organic chemistry of cells well enough to treat cells like complex machines -- to really do cellular engineering."
All cell surfaces are decorated with oligosaccharides -- complex structures strung together inside the cell from a few simple sugars. Different kinds of cells display different oligosaccharides, and even the same kinds of cells display different patterns depending on their stage of development or environment. Since each oligosaccharide is chemically unique, each imparts to the cell a unique surface for interaction with the outside world.
"We asked ourselves, how can we exploit these differences?" says Bertozzi. Working with graduate student Lara Mahal and postdoctoral fellow Kevin Yarema, Bertozzi set out to design new cell surfaces that could stick to synthetic materials. "We decided to appropriate the cell's natural metabolic machinery for assembling tailor-made oligosaccharides."
Bertozzi reasoned that if a properly designed synthetic sugar with novel chemical properties could be ingested by the cell, the sugar might be incorporated in an oligosaccharide and delivered to the surface. The result would be a cell with new surface properties.
To demonstrate the technique, she and her colleagues chose an analogue of sialic acid, a sugar which in its natural form is often found in the cell-surface oligosaccharides of human cells. "We planned to use an unnatural sugar related to sialic acid, one that carries an unnatural functional group. We hoped that if the cells ate the unnatural sugar -- without noticing, so to speak -- they would install it along with its functional group in oligosaccharides, and thus decorate themselves with these unnatural markers."
To tag the sialic acid, Bertozzi's team needed a functional group that wasn't normally found on cell surfaces but wasn't harmful either, one that could react with other groups on synthetic materials -- and under physiological conditions, such as a watery environment and mammalian body temperature.
They chose the ketone group. Rarely found on cell surfaces, ketones react strongly with a functional group called the hydrazide; the special reactivity of the ketone could allow a selective affinity for materials that had been outfitted with the hydrazide group, such as ceramics, organic thin films, and metals.
The natural chemical precursor of sialic acid is called N-acetyl mannosamine -- more conveniently known as ManNAc -- but Bertozzi and her colleagues fed cultured cells an artificially synthesized precursor known as ManLev, identical except that it contains a ketone group. The cells consequently manufactured sialic-acid oligosaccharides with ketones and expressed them in copious amounts on their surfaces -- over a million copies on the surfaces of most cells. Moreover, the researchers found they could precisely control the degree of ketone labeling by adjusting the relative amounts of natural (ManNAc) and unnatural (ManLev) precursors fed to the cells.
Cell surfaces modified for specific reactions hold great promise in the construction of biocompatible materials and artificial organs, but the interests of Bertozzi's cell-surface group don't stop there. "We're investigating the cells of other organisms, such as plants and microbes. We're looking into biosensors, in which cells designed to lock onto specific compounds can be combined with an electronic, transducing substrate to signal changes in the environment" -- a sort of cyborg canary-in-a-coal-mine.
Another possible use of reactive chemical groups on cells emerged while Bertozzi, Mahal, and Yarema were developing the technology. "Many human cancer cells, including colon, breast, and prostate cancers and certain leukemias, have aberrant patterns of oligosaccharides," says Bertozzi. "For one thing, they show extremely high levels of sialic acid. The possibilities were obvious."
Bertozzi and her colleagues showed that ketone-labeled cancer cells, otherwise robust, could be made uniquely vulnerable to a derivative of the natural plant toxin ricin. The ricin analog, synthetically armed with the reactive hydrazide group, sought out and reacted with the ketone-labeled cells.
"It worked," says Bertozzi. "We killed 'em."
Bertozzi's group is currently moving studies of hydrazide-labeled toxins from the test tube into laboratory animals. Other labeled cancer-killers are being explored, as well as a method for making ketone-labeled cancer cells stand out in magnetic-resonance imaging by using hydrazide-labeled compounds for high contrast.
Bertozzi's team has thus become the first research group to install specific functional groups on the cell surface through metabolic mechanisms; the tools they used were "an equal combination of cell biology and synthetic organic chemistry," says Bertozzi, who intends her cell-engineering methods to be simple and rational enough to be understood and used by biologists and chemists working together.
Bertozzi, Mahal, and Yarema have recently written about cell surface engineering in Science, 16 May 1997, vol 276, page 1125, and in the June, 1997 issue of Chemistry & Biology, volume 4, page 415.
The Berkeley Lab is a U.S. Department of Energy national laboratory located in Berkeley, California. It conducts unclassified scientific research and is managed by the University of California.
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