Medical Device & Diagnostic Industry MagazineMDDI Article IndexOriginally Published March 2000R&D HORIZONSEfforts to develop viable alternatives to human blood are beginning to prove successful in both operating suites and emergency rooms.

Gregg Nighswonger

March 1, 2000

18 Min Read
Blood Substitutes: Recreating the Fluid of Life

Medical Device & Diagnostic Industry Magazine
MDDI Article Index

Originally Published March 2000

Efforts to develop viable alternatives to human blood are beginning to prove successful in both operating suites and emergency rooms.

In addition to being a critical factor influencing the recovery of patients with severe blood loss through trauma or surgery, blood is an essential element in effectively treating a number of other injuries and conditions, including shock. Additionally, transfusions with whole blood and blood components are widely used as part of the therapeutic regimen in treating a broad range of illnesses and conditions, including anemia resulting from various sources, cancer and leukemia, sickle cell disease, and other disorders. Some estimates suggest that within a few years, demand for red blood cells in the United States is likely to be divided among three major categories. About 55% will continue to be required for elective and general surgical procedures. Another 30% of red blood cells will be used for patients with chronic or acute anemia, and certain other blood disorders. An estimated 15% will be used for trauma and emergency procedures.

According to the American Association of Blood Banks, an estimated 4 million patients in the United States each year receive transfusions of whole blood, including an estimated 3 million surgery patients; approximately 12.6 million units are donated and about 643,000 are autologous. Additionally, more than 23 million units of blood components are transfused each year.

At a fraction of the size of human red blood cells, hemoglobin-based oxygen carriers can be effective in reducing the need for donor blood during surgery.

A number of factors drive the need for a viable substitute for human blood—including the need to eliminate transfusion-related transmission of infectious disease, reduce the need for cross-matching and related costs, and increase shelf life and stability at ambient temperatures. Current transfusion-related risks, including infection and hemolytic reactions, are generally considered to be central to the need to develop a blood substitute. Minor reactions to transfusions are on the order of 1 adverse reaction per 100 transfusions, or 1.0%; the probability of a fatal hemolytic reaction is estimated to be 0.001%. Transfusion-related infection risks are estimated to be 0.002% for hepatitis B, and 0.03% for non-A non-B hepatitis. The chance of becoming infected with HIV as the result of a transfusion is currently 1 in 225,000.


It is generally possible to survive very low hematocrit levels (red blood cell volume as a percentage of blood volume) resulting from losses of up to 70% of red blood cell mass; however, the body's ability to compensate for much smaller losses of blood volume is quite limited. A loss of only 30% of blood volume can lead to irreversible shock if not treated rapidly enough. Various fluids have been found to be appropriate for increasing the blood volume, and are classified as crystalloid solutions, colloidal solutions, or oxygen-carrying solutions. Crystalloid solutions, such as Ringer's lactate or saline, and colloidal solutions, including use of albumin as a plasma expander, have proven valuable as a blood substitute on the basis of volume replacement. Oxygen-carrying solutions, on the other hand, have posed much greater challenges, and the object of most research efforts has not been to find a replacement for whole blood, but to duplicate its oxygen-carrying capabilities.

In most situations, patients requiring blood replacement because of blood loss need only a short-term replenishment of the oxygen-carrying capacity of hemoglobin until their own bodies synthesize replacement red blood cells. It is the hemoglobin, however, that requires refrigeration, has a relatively short shelf life, and must be carefully matched for correct blood type and other factors. Efforts to develop a viable blood substitute have thus focused on creating a hemoglobin alternative that can be stored for a long period of time at room temperature and can be transfused to restore the oxygen-carrying function of hemoglobin without the need for type matching. According to FDA estimates, at least 11 companies have blood substitute products publicly disclosed and under development in preclinical and end stages of clinical trials. By the beginning of 1999, four different hemoglobin-based oxygen carriers (HBOCs), developed by four separate companies, had entered the late stages of clinical development.

A problem to be addressed in working with hemoglobin itself is that its behavior changes with separation from the red blood cells. Once separated, it divides into halves that are no longer capable of oxygenating tissue, and which can cause kidney damage. Researchers have succeeded, however, in overcoming this challenge to administering free hemoglobin by developing a cross-binding reagent that is capable of preventing the hemoglobin molecules from splitting after removal from the red blood cells.

Attempts to develop blood substitutes have followed a number of different strategies. There has been some success with developing methods for extracting and chemically processing hemoglobin from donated human blood. Although this method can address the challenge of creating a whole blood substitute that requires no typing and is more stable, it still relies on the availability of donated blood. Methods for genetically engineering hemoglobin that remains stable outside red blood cells and are capable of releasing adequate amounts of oxygen without a cross-binding reagent are seeing some success. Researchers working in this area are working with a mutant gene form that produces a modified form of hemoglobin with the proper characteristics. Once the gene is inserted into bacteria, the desired hemoglobin is produced. Genetic engineering has also been used to create transgenic pigs capable of carrying the gene for producing normal human hemoglobin while retaining the gene for producing swine hemoglobin. Hemoglobin is harvested from blood drawn harmlessly from the pigs, then ruptured and the human hemoglobin separated out. The separated hemoglobin can be processed to remove any infectious agents. Similar methods are being explored to produce a blood substitute based on bovine hemoglobin.


An Ohio firm specializing in perfluorocarbon technologies, Synthetic Blood International (SYBD; Kettering, OH), is developing a blood substitute based on that technology. Because blood gases such as oxygen and carbon dioxide are highly soluble in perfluorocarbons, SYBD's Oxycyte is intended to provide an effective means of transporting oxygen to tissues and carbon dioxide to the lungs. Compared with hemoglobin, Oxycyte has been found to be capable of carrying at least five times more oxygen. Additionally, perfluorocarbons are considered to be more effective than hemoglobin for delivering oxygen at the tissue level. According to the company, the perfluorocarbon microdroplets that carry the oxygen are 1/70th the size of the red cells. They can therefore reach many areas of the body that human red blood cells cannot.

The product is inert and can be fully sterilized. It can be stored at room temperature and does not require typing and cross-matching prior to use. The firm considers Oxycyte to be an all-purpose synthetic blood product, with potential uses including surgery, trauma, angioplasty, open heart surgery, and oxygenation of tumors during radiation or chemotherapy. The product can also be made available on the battlefield, at the scene of accidents, and stored in emergency vehicles and emergency departments.

SYBD notes that its consultant director of research, Leland C. Clark Jr., PhD, pioneered the application of biocompatible, oxygen-carrying fluorocarbons, and the incorporation of these into synthetic blood emulsions for intravenous use more than 20 years ago. Clark's research formed the basis for commercialization of a synthetic blood emulsion in Japan, and of a perfluorocarbon emulsion, Fluosol, in the United States for use with percutaneous transluminal coronary angioplasty procedures.

Another firm, Alliance Pharmaceutical Corp. (San Diego), is developing Oxygent as a perfluorocarbon-based blood substitute to temporarily augment oxygen delivery in patients at risk of acute tissue oxygen deficit due to transient anemia, blood loss, or ischemia. According to the company, Oxygent, which is in phase-3 clinical development, is a stable, concentrated emulsion of tiny particles, about 0.2 µm in diameter, suspended in a water-based solution. The particles have a perfluorochemical core consisting primarily of perflubron surrounded by a surfactant derived from egg yolk lecithin.

Alliance notes that Oxygent is used according to the firm's proprietary augmented acute normovolemic hemodilution (Augmented-ANH) method. Like other methods described in this article, the method is designed to enhance conservation of the patient's own blood, minimizing surgical blood loss. "We expect that Augmented-ANH with Oxygent will be the most cost-effective proposed alternative to donor blood for surgical patients," says Alliance chairman and CEO Duane J. Roth. The company states that Oxygent has been administered to patients in clinical trials involving more than 850 subjects. Two phase-3 studies with cardiac and general surgery patients are ongoing in the United States and eight European countries.

Synthocytes, a platelet substitute, are being developed to treat thrombocytopenia, a common result of chemotherapy.

Also focusing on perfluorocarbon-based products, Sanguine Corp. (Pasadena, CA), recently sent a letter to its shareholders indicating that it was planning to begin animal trials of a second-generation perfluorocarbon-based artificial blood based on the PHER-02 molecule. The product can be administered at room temperature, has a shelf life of three years, and requires no typing or cross matching, according to the firm. The Sanguine molecule is approximately 1/900th the size of human red blood cells, yet can deliver three to four times the amount of oxygen, and can remove carbon dioxide similarly to red blood cells. Because of the molecule's small size, PHER-02 is capable of bypassing vascular obstructions to deliver blood to occluded vessels, restoring and maintaining tissue viability. This is a capability not typically found in blood substitutes, according to Sanguine. The firm states that the product provides certain advantages for treating victims of shock or ischemia, and that its long shelf life and ability to be used without typing make it useful when disasters, trauma, and accidents require immediate oxygenated blood replacement.


Biopure Corp. (Cambridge, MA) recently announced results of clinical studies involving its investigational oxygen therapeutic, Hemopure, a bovine-derived HBOC product. The results suggest that the product can eliminate the need for allogeneic red blood cell transfusions in a significant number of patients undergoing vascular surgery. According to Glenn M. LaMuraglia, MD, the principal investigator at the division of vascular surgery, Massachusetts General Hospital, one of eight hospitals that participated in the study, "This oxygen therapeutic solution eliminated intra- and postoperative red blood cell transfusions in more than a quarter of these high-risk patients despite study design restrictions on the total dose and number of days over which the product could be administered."

Specifically, the results indicated that the blood substitute "totally eliminated the need for red blood cell transfusions in 27% of patients throughout the entire 28-day follow-up period," according to the firm. "Moreover, 39% of Hemopure patients did not require allogeneic blood during the 96-hour treatment period and 66% did not require allogeneic blood on the day of surgery, further supporting the product's use as an oxygen bridge." Hematocrit levels were similar for both the Hemopure group of patients and the control group at the time of discharge from the hospital. The company believes that Hemopure may support the production of the patient's own red blood cells.

To date, Hemopure has been administered to more than 600 patients in 21 completed or ongoing clinical trials. One unit of Hemopure contains 30 g of ultrapurified, chemically cross-linked hemoglobin in 250 ml of a balanced salt solution. When infused, this linked hemoglobin circulates in the plasma, and has a lower viscosity and more readily releases oxygen to tissues than to blood. Hemopure has been shown to be stable at room temperature for at least 30 months. The product is compatible with all blood types, and is purified through patented techniques that are validated to remove infectious agents, including bacteria, viruses, prions, and other potential contaminants, the company states.


Earlier this year, Hemosol Inc. (Toronto) announced positive results from a clinical trial of its Hemosol hemoglobin replacement product involving 60 patients undergoing coronary artery bypass grafting (CABG) procedures. Hemolink is an HBOC derived from human red blood cells. Used in surgical applications, the product is used in conjunction with intraoperative autologous donation (IAD). IAD is a blood conservation strategy in which up to four units of a patient's blood are removed just prior to undergoing surgery. This harvested blood volume is replaced with Hemolink instead of conventional volume expanders that do not deliver oxygen to tissue. The patient's harvested blood is subsequently returned as needed, as well as blood recovered by cell salvage methods. The result is a reduction or complete elimination of patient exposure to donated blood.

The company believes that use of Hemolink can provide "greater assurance of safety from viral and bacterial transmission, a reduced risk of allergic or immune reaction, universal compatibility with all blood types, efficient oxygen delivery to vital organs and tissue, and an extended shelf life of at least one year.

Conducted at nine U.S. centers, the Hemolink study specifically involved CABG patients who were at increased risk of blood loss and would require transfusion with donor blood. The object was to determine whether Hemosol's HBOC product "could be safely given to patients undergoing this type of surgery, and whether the use of donated blood could be either significantly reduced or avoided compared with a best available treatment control arm," according to the company. The results suggest that the product was safe and resulted in no clinically limiting side effects.

Similar results were reported in clinical studies conducted in Canada and the United Kingdom. In these earlier studies, involving 60 patients with moderate blood loss, 90% of patients receiving at least 750 ml of Hemolink avoided donor blood, compared with approximately 55% of patients receiving the best available treatment in the control arm. Studies are also being conducted involving orthopedic surgery, anemia, and other cardiovascular procedures, the firm states.

Another chemically modified hemoglobin product, PolyHeme, has been developed by Northfield Labs Inc. (Evanston, IL) and has shown positive results in clinical studies. The product uses a solution of donated human blood purchased from the American Red Cross and Blood Centers of America for use as a starting material. A proprietary process is used to separate, filter, and chemically modify the blood to produce PolyHeme. Hemoglobin extracted from the red blood cells is filtered to remove impurities. A multistep process is then used on the purified hemoglobin to chemically modify it, creating a polymerized form of hemoglobin designed to avoid the undesirable effects typically associated with hemoglobin-based blood substitutes. The modified hemoglobin is then incorporated into a solution that can be administered as an alternative to transfused blood. According to Northfield, a single unit of PolyHeme contains 50 g of modified hemoglobin—about the same amount of hemoglobin delivered by one unit of transfused blood.

In the first randomized, prospective trial of PolyHeme, surgeons in three university-based trauma centers obtained informed consent to enroll 44 severely injured trauma patients. Of those, 23 were randomized to receive donated red cells as their initial blood replacement, and 21 to be given the polymerized hemoglobin product.

According to the researchers, the blood substitute was found "at least as physiologically active as red cells in loading and unloading oxygen." No serious or unexpected adverse events were reported in patients who received the blood substitute. Assessments of patient organ function were unchanged except for a slight elevation in total bilirubin. The researchers further note that the substitute reduced the amount of donated blood these patients subsequently needed by nearly half—6.8 additional units, compared with 10.4 additional units required by patients initially treated with donated red cells. The company states that the results bring researchers closer to a resuscitative fluid that is "universally compatible, immediately available, free of vasoactive properties, free of disease transmission, and capable of long-term storage."

In another study involving 53 patients who received between 6 and 20 units of the blood substitute, none of the patients received banked blood during the study period, according to the company. Of the 53 patients, 27 had less than 25% of their own blood after infusion with PolyHeme and, of those 27, 20 had less than 12% of their own blood remaining. Five had less than 6% of their blood remaining after the infusion. Northfield indicates that the 27 patients with less than 25% of their own blood would have had an expected short-term survival rate of less than 20%, if banked blood or a blood substitute had not been available; however, 85% of the patients survived after infusions of PolyHeme. The firm suggests that this is essentially the same percentage that would have been expected to survive if banked blood had been used.

Northfield believes that, because the survival rates mirrored those for patients receiving blood, these results demonstrate the product's ability to provide an enhanced survival rate for bleeding patients in situations where blood might not be available. The company states that, "PolyHeme is the only blood substitute undergoing clinical trials that has been tested at large enough dosages to be considered a substitute for acute blood loss in trauma and surgical settings." Although the data from these initial studies are promising, they are still considered small and much larger studies are believed to be needed.


In addition to efforts to find blood substitutes, research focusing on platelet substitution is being driven by potential use in treating certain conditions that are common among cancer patients. One of the side effects of cancer therapy is the drastic reduction in platelets, or thrombocytopenia. The condition is currently treated with a transfusion of blood-derived platelets. As of 1998, it was estimated that 18 million units of platelets are transfused each year worldwide, 80% of which are given to patients who are thrombocytopenic as a result of chemotherapy. As chemotherapy regimes become more and more aggressive and as the use of bone marrow transplantation increases, the requirement for platelets will grow.

Among the products being considered specifically to treat thrombocytopenia are Synthocytes, in development by Andaris Group Ltd., a subsidiary of Quadrant Healthcare plc. (Nottingham, UK). Synthocytes are microcapsules to which fibrinogen has been chemically linked. Acting as a replacement for human blood platelets in the prevention of bleeding, the Synthocytes are capable of selectively targeting the site of hemorrhage, according to the company. The product is believed to offer certain key advantages over blood-derived platelets, which have the potential to transmit viral infections, suffer from instability during storage, and cause immune reactions.


Research into development of blood substitutes is continuing at numerous centers. At the University of Pittsburgh medical center's McGowan Center for Artificial Organ Development (Pittsburgh), researchers are focusing on artificial blood components that can function as an all-purpose blood substitute. The McGowan artificial blood components are being designed to combine the high oxygen-carrying and nutrient supplement capability of natural blood with superior fluid properties. According to the researchers, "The significant novelty of our research is to achieve maximal oxygen delivery to tissues with minimal concentration of oxygen carrier via the superior mechanical and physical properties of the product. Additional advantages of the product are low toxicity and a low cost due to significantly reduced concentration of the oxygen carrier." The McGowan artificial blood components are being tested in vitro for their effectiveness and stability. The first phase of the functionality, toxicity, and general biocompatibility testing preparations is still in progress, and the patent application on the novel artificial blood was filed in September 1998. The researchers note that one possible application of artificial blood is protection of blood cells from mechanical damage in blood-wetting artificial organs.


Efforts to develop artificial blood products are global in nature. Extensive research has been conducted in Canada, Europe, and Asia. In May of 1999, for example, an announcement was made during a meeting of the Royal Australasian College of Surgeons that a new blood substitute, polymerized hemoglobin, would be widely available to surgeons in Australia and New Zealand within one to two years. The announcement was made by Ian Civil, head of the RACS Early Management of Severe Trauma program, who explained that polymerized hemoglobin differs from blood products designed to carry oxygen throughout the body in that it poses fewer dangerous side effects, such as suppressing the patient's immune system. The product is also said to eliminate the need for cross-matching blood types. In addition, the product can be stored without refrigeration. According to Civil, "This means that, if we have a problem with a patient bleeding, instead of having to cross-match the patient with the donor and run the risk—albeit small—of transferring blood-borne diseases, we can just reach up to a shelf and use polymerized hemoglobin." Polymerized hemoglobin has been used in treatment of trauma patients in the United States and is undergoing clinical trials for FDA approval.


Although notable progress has been made in bringing effective replacements for whole blood into clinical practice, most experts still consider artificial blood to be at or near the frontiers of development. Non-oxygen-carrying substitutes for human blood, such as Ringer's lactate and other colloidal solutions, currently provide an effective treatment for blood losses on the order of 50% with few if any negative side effects.

Replacement of human blood with an oxygen-bearing substitute is considered essential for extreme blood loss. Although oxygen-carrying replacements based on perfluorocarbons have demonstrated numerous positive features, their use has been associated with certain limiting side effects. HBOCs capable of effectively treating hypovolemia are intrinsically capable of providing oxygen for tissues, yet the feasibility of successfully introducing large volumes of a hemoglobin-based substitute to the body must be examined further. Microencapsulation of modified hemo-globin may further enhance the use of HBOCs. Additional factors include an increasing need for blood, potential risks associated with donated blood (e.g., infection, allergic reaction, and hemolytic reaction), and the potential benefits of a substitute offering extended shelf life. Ultimately, the use of blood substitutes may require that a fine balance be drawn between the need for its life-giving properties and the potential severity of its side effects.

Gregg Nighswonger is executive editor of MD&DI.

Illustration by MICHAEL HIRANO

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