May 1, 2011 (Vol. 31, No. 9)

Editor’s Note

With a number of monoclonal antibody products already on the market and many others in clinical trials, it’s clear that mAbs play an essential role in diagnosing and treating various diseases and medical disorders in the 21st century. But smaller genetically engineered antibody fragments (e.g., Fab, scFv) are catching the eye of a growing legion of scientists, since the fragments demonstrate the same targeting capabilities as whole monoclonals. They also can be produced less expensively.

To help commemorate GEN’s 30th anniversary in 2011, we’ve reprinted an article from the July/August 1988 issue of GEN that marks a milestone in the production of these promising Fab molecules. The GEN article reports that researchers at International Genetic Engineering succeeded in engineering the first bacterium, E. coli, to produce Fab proteins after previously carrying out the same study in yeast cells.

The scientists interviewed in the article noted that the new methodology should cut costs and be more efficient than hybridoma methods being used at the time. The other goal was to engineer future Fabs for different functions. This and similar developments over the past 20 or so years set the stage for current R&D projects focusing on Fabs.

—John Sterling, Editor in Chief

“As Seen in GEN—Flashback” Volume 8, No.7, July/August 1988

Researchers Produce Fab Proteins in E.coli

By Judy Berlfein

Scientists at International Genetic Engineering, Inc. (Ingene) of Santa Monica, Calif. have engineered the first E. coli to produce Fab proteins. The antibody fragments bind specifically to a human colon cancer cell line. Earlier this year, the company announced they were also synthesizing Fab proteins in yeast cells.

“There’s a real need for a quick way of making Fabs,” Dr. Ivor Royston, director of clinical immunology at the University of California San Diego Cancer Center said. “It looks like Fabs are going to be very useful for imaging cancers.”

The fragments penetrate tumors faster than whole antibody which is more advantageous when using isotopes with short half-lives.

For example, technetium and iodine-123 both have half lives of approximately 11 or 12 hours. Whole antibody takes two to three days to penetrate a tumor and by then the isotope has lost activity and produces a poor image. With a Fab fragment, the antibody enters the tumor quickly while the isotope is still intact. In addition, Dr. Royston added, there is evidence that by using Fabs, there may be less non-specific uptake in the liver.

Less Costly

Dr. Marc Better, project scientist at Ingene and first author of the paper which appeared in Science, claimed the new technique should cut costs and be more efficient than present hybridoma methods. Bacteria reproduce in 20 minutes whereas mammalian cells require at least a day. With hybridomas, Fab proteins go through two purification steps—first with whole antibody and then with papain-cleaved fragments. E. coli, however, produces Fabs directly and the proteins are only purified once.

In addition, media and equipment cost less for bacterial fermentation than for mammalian tissue culture. “As long as you have bacteria that are producing a reasonable quantity of these materials,” Dr. Better noted, “then producing them in bacteria will be much less expensive.”

Using E. coli, one synthesizes uniform Fab proteins. With papain, it is common to create a mixture of varied antibody fragments. “When people try to crystallize Fabs, they often run into trouble because they don’t have a homogeneous preparation,” Dr. Better emphasized. Now that genetically engineered antibodies are available, this problem will be alleviated.

First Step

This is only the first step however. Future Fabs may be engineered specifically for different functions. “We may be able to make a whole army of slightly different molecules which may have more desirable properties,” Dr. Better said. They may design a Fab with reduced immunogenicity or introduce a binding site for a therapeutic agent.

But the new technology won’t be putting the hybridoma industry out of business. Whole antibodies still have an important function. And according to Dr. Better, E. coli may never be able to produce these highly complex molecules.

Who will take over the Fab market—the genetic engineers or those using papain-cleaved antibody? “My best guess,” Dr. Better replied, “is that whoever can make the reagents the cheapest is going to win.”

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