In Search of a Substitute for Life’s Elixir : Medicine: For 300 years, physicians have sought a fluid that could replace blood in emergencies. At last, researchers may be close.
Physicians began to search in the 17th Century for potent fluids that could substitute for blood. They experimented by transfusing wine and ale into animals; they tried using dog and lamb blood in human patients--with fatal results. In a bizarre 19th-Century fad that swept the United States, medical researchers even infused milk from cows, goats and humans into the veins of patients.
Today’s genetic engineers are also using methods that might strike future generations as bizarre: manipulating the genes of pigs to produce human blood cells, brewing batches of blood molecules in otherwise lethal bacteria, designing the structure of synthetic blood compounds on a computer screen.
But with heightened concerns about the health risks of blood transfusions, biotechnology firms and government research groups are racing to find a plentiful and safe alternative to the lifesaving elixir.
At least two products are being tested in human studies, although scientists are skeptical that artificial blood will soon be on pharmacy shelves. The AIDS epidemic has given their work a new sense of purpose to their work, even though the risk of contracting the AIDS virus in blood transfusions now is minimal, about one in 225,000 units of transfused blood.
As Robert Winslow, a professor of medicine at UC San Diego and former chief of the blood research division of the Army’s Letterman Institute of Research in San Francisco, put it: “In the early 1980s, people realized we need an alternative to red blood cells because of the issue of AIDS.”
But making blood is not simple. In the search for a substitute, scientists long ago gave up the dream of precisely replicating the complex, sticky red formulation of whole blood. “It defies laboratory synthesis,” wrote William R. Amberson, a physician at the University of Maryland School of Medicine, in 1937.
Instead, scientists have been targeting only one component of blood: the oxygen-carrying protein molecule called hemoglobin, which resides inside the protective membrane of the red blood cell. Hemoglobin is the magically restorative part of blood that carries oxygen to all parts of the body. Without oxygen, the tissues die.
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The military also has an obvious interest in blood substitutes. “Traditionally, the Army has always supplied blood products for . . . combat troops from blood drawn in the United States,” said Gerald Moore, assistant chief of the blood research division of the Letterman Institute.
“Shipping it around the world and storing it has always been a major problem,” one that could be solved by a substitute with a long shelf-life. “Our dream some time back was to have this material freeze-dried, like coffee: add water, shake and use,” he said. “It’s still a dream.”
Although scientists have been experimenting with hemoglobin since 1916, they are far from devising an ideal product. Blood substitutes under development today have at best a circulation time of 24 hours before they are metabolized by the body, making them only temporary stand-ins until donated human blood can be transfused. Regular blood from transfusions remains effective in the circulation an average of 34 days.
But lowered expectations may lead to a more realistic chance of developing a useful blood substitute, many researchers say. “It is better to think of these solutions as resuscitative fluids,” said Steven Holtzman, executive vice president of DNX, a biotechnology company in Princeton, N.J.
At present, only two companies, Baxter Healthcare Corp. and Somatogen Inc., have hemoglobin substitutes in early clinical trials in which new products are tested on people with the approval of the Food and Drug Administration. Moore said he expects that approval of any product for general use is at least five years away. “And we’ve been saying this for 10 years,” he added.
What presents the greatest challenge is the large therapeutic dose. Most drugs are given in small quantities, but hemoglobin substitutes would be administered in pints, greatly amplifying any bad effects.
“This is a real adventure that’s unparalleled,” said Winslow. “Some of the materials being studied today don’t stay in the blood vessels where you want them to. Blood comes in contact with every tissue in the body. The possible consequences and side effects are enormous.”
Companies making a hemoglobin blood substitute do not have any problem proving its effectiveness, a task faced by makers of most other new drug products. “We can show it carries oxygen and aids in the resuscitation of trauma patients,” said Thomas Schmitz of Baxter Healthcare Corp. in Round Lake, Ill. “The real issue is safety. . . .”
Before current efforts, hemoglobin substitutes have always had toxic and potentially lethal side effects when tested in humans.
“There have been 12 or 15 published clinical trials done in the last 20 years, and a lot of these people got sick,” said Moore. “Every time we peel away a layer, we find another problem underneath. We are just now getting down to the fundamentals of the toxicity.”
Because red blood cells cannot be made disease-free through sterilization without being destroyed, researchers for 30 years have been working with hemoglobin taken out of the cell. So-called “cell-free hemoglobin” can be heat-treated and chemically treated without its oxygen-carrying capability being destroyed. Moreover, hemoglobin outside of the cell membrane does not tend to provoke an immune response.
Until recently, outdated donated human blood was the source for cell-free hemoglobin experiments. Now, some companies are genetically engineering animals and bacteria to produce human hemoglobin. But it has not yet been proven that hemoglobin removed from its cell can be made safe for human beings, regardless of the source.
The problem has been that, free of the cell, hemoglobin actually binds too tightly to oxygen and won’t give it up to the tissues. Also, the free hemoglobin molecule is unstable and tends to break up into smaller molecules, which can cause a toxic blockage of the kidneys. In earlier experiments, patients have reacted with fevers, dangerously high blood pressure, chest pain and flu-like symptoms, including nausea.
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To try to make cell-free hemoglobin safe, biotech companies have taken two basic approaches. One is to chemically modify the structure of the hemoglobin so that it doesn’t hold so tightly to oxygen and will not break up into small particles that could get caught in the kidneys.
The second approach is to do the modification at the genetic level, before the hemoglobin is produced. That way each cell contains the change.
Somatogen, in Boulder, Colo., takes the genetic approach. The company uses genetically engineered e. coli bacteria, outfitted with human DNA, to “brew” human hemoglobin in large vats.
This bacteria is normally deadly but is used as a kind of factory to produce desired proteins because the bacteria reproduce so quickly. The proteins produced--in this case human hemoglobin--are not poisonous and can be chemically cleaned of any toxic bacterial residue. In Somatogen’s laboratories, the genetic blueprint instructing the e. coli bacteria to produce the hemoglobin was modified to make a more stable structure than natural cell-free hemoglobin.
“When you’re thinking about how to design a drug (with the computer), you can manipulate the structure right in front of you,” said Dave Anderson, director of protein engineering for Somatogen. “We can very easily make changes in the hemoglobin on the screen. It’s similar to an engineer looking at a blueprint for a bridge.”
Charles H. Scoggin, Somatogen’s president, said the company has come up with about 100 different versions of cell-free hemoglobin. “We cut and paste within this gene to make new forms of hemoglobin,” said Scoggin.
Somatogen’s latest version, called rHb1.1, has been administered to healthy volunteers in a single dose of up to 11.5 grams, about half the therapeutic dose. “There were no serious adverse events,” said Steven Shoemaker, senior vice president. “The only side effects we saw were transient elevations of temperature, some headaches, muscle aches and nausea, and these either resolved spontaneously or after treatment with ibuprofen.” The FDA has given Somatogen permission to test dosages up to 25 grams.
In another approach, DNX scientists have created a line of genetically engineered pigs that produce human hemoglobin as a portion of their blood, usually from 10% to 15%.
The pig-generated hemoglobin would then be chemically modified to eliminate safety problems. Scientists would also have to purify the pig-generated blood to prevent contamination with either animal or human viruses. “We are investigating various methods of virus removal and deactivation,” Holtzman said. “Because swine are not known to be susceptible to AIDS or hepatitis, we don’t have the same concern about those particularly human pathogens.”
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Last July, Baxter Healthcare Corp. began human studies of a chemically modified hemoglobin product derived from outdated human blood collected from voluntary donors.
“Our raw material is probably inherently relatively safe,” said Schmitz. “There are no foreign proteins or foreign substances. And the level of purification we need to do is probably less demanding than for hemoglobin from other sources.”
Baxter is at an interim point in the first phase of a safety study and hopes to finish the study by year’s end, said Schmitz.
Northfield Laboratories Inc. conducted clinical trials in 1989 of a chemically modified hemoglobin product but suspended the trials after subjects complained of chest pains.
Upjohn began clinical safety trials in mid-1991 of a blood substitute made by Biopure Corp. of Boston that was derived from purified cattle blood. But Upjohn quickly stopped the trials after an undisclosed problem. The company is waiting for FDA permission to begin testing a new version of Biopure’s bovine hemoglobin molecule.
Carl W. Rausch, a chemical engineer who is chairman and a co-founder of Biopure, said that there had been no allergic reactions to the first version but that some patients had experienced “a transient medical event that resolved itself within 12 hours.”
“Once you get over the hurdle of an immune response,” he said, “the whole issue becomes one of purification and availability.” He said Biopure was using bovine blood because of its vast supply from cattle slaughtered for food.
Rausch said he believes that the problems with hemoglobin substitutes tested on humans in the past were probably a result of insufficient purification.
“The purification techniques just weren’t available prior to the mid-’80s,” he said. “Knowing that, all hemoglobin work before 1985 or 1986 has got to be suspect.” Schmitz also ascribes past failures to improper manufacturing techniques. “Obviously, we think that’s a very strong possibility,” he said.
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Can cell-free hemoglobin be made safe enough? Which manufacturing method will prove the best? “We are just now starting the studies that will answer those questions,” said Schmitz. “They will certainly get answered in the next two years.”
Other scientists are more wary. Somatogen’s genetic approach to altering hemoglobin “is sophisticated and scientifically pleasing,” said Harvey G. Klein, a scientific adviser to the company and chief of the department of transfusion medicine at the National Institute of Health’s clinical center. “But whether it will turn out to be better (than chemically modified hemoglobin) is still a matter of scientific conjecture.
“The multimillion-dollar question on this--it used to be the $64 question--is hemoglobin taken outside of the red cell innately toxic?” he said. “It could be the Good Lord put it into little red-cell packets for a reason. The hemoglobin may cause problems we don’t know.”
Moore said the Army keeps running into safety problems. A hemoglobin substitute “is doable,” he said, “but it will not be without potential side effects. It won’t be a totally benign solution.”
Winslow believes that in the end, cell-free hemoglobin may have to be re-enclosed in some kind of neutral envelope before it can be used safely in large quantities. But he is optimistic that a red-cell substitute will be found.
Compared to earlier attempts to make artificial blood, he said: “What’s different now is that many more good and serious people are getting into the research. More and more resources are being focused on this problem.”
