Blood substitutes

Blood substitutes, often called artificial blood, are used to fill fluid volume and/or carry oxygen and other blood gases in the cardiovascular system. Although commonly used, the term is not accurate since human blood performs many important functions. Red blood cells transport oxygen, white blood cells defend against disease, platelets promote clotting, and plasma proteins provide various functions. The preferred and more accurate terms are volume expanders for inert products, and oxygen therapeutics for oxygen-carrying products. Examples of these two "blood substitute" categories:

• Volume expanders: inert and merely increase blood volume. These may be crystalloid-based (Ringer's lactate, normal saline, D5W (dextrose 5% in water) or colloid-based (Haemaccel, Gelofusin).

• Oxygen therapeutics: mimic human blood's oxygen transport ability. Examples: Hemopure, Oxygent, PolyHeme.

Oxygen therapeutics are in turn broken into two categories based on transport mechanism: perfluorocarbon based, and hemoglobin based.

Volume expanders are widely available and are used in both hospitals and first response situations by paramedics and emergency medical technicians. Oxygen therapeutics are in clinical trials in the U.S. and Europe, however Hemopure is more widely available in South Africa.

Artificial blood is also often used in movies.

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Volume expanders

When blood is lost, the greatest immediate need is to stop blood loss. The second greatest need is replacing the lost volume. This way remaining red blood cells can still oxygenate body tissue. Normal human blood has a significant excess oxygen transport capability, only used in cases of great physical exertion. Provided blood volume is maintained by volume expanders, a quiescent patient can safely tolerate very low hemoglobin levels, less than 1/3rd of a healthy person.

The body automatically detects the lower hemoglobin level and compensatory mechanisms start up. The heart pumps more blood with each beat. Since the lost blood was replaced with a suitable fluid, the now diluted blood flows more easily, even in the small vessels. As a result of chemical changes, more oxygen is released to the tissues. These adaptations are so effective that if only half of the red cells remain, oxygen delivery may still be about 75 percent of normal. A patient at rest uses only 25 percent of the oxygen available in his blood. In extreme cases, patients have survived with a hemoglobin level of 2 g/dl, about 1/7th of normal, although levels this low are very dangerous.

With enough blood loss, ultimately red cell levels drop too low for adequate tissue oxygenation, even if volume expanders maintain circulatory volume. In these situations the only alternatives are blood transfusions, packed red cells, or oxygen therapeutics (if available). However in some circumstances hyperbaric oxygen therapy can maintain adequate tissue oxygenation even if red cell levels are below normal life sustaining levels.

Towards artificial blood

Artificial blood is supposed to fulfill some functions of biological blood, especially in humans. The term oxygen therapeutic is more accurate, as human blood performs other functions besides carrying oxygen. For example white blood cells defend against infectious disease, and platelets are involved in blood clotting.

The initial goal of oxygen carrying blood substitutes is merely to mimic blood's oxygen transport capacity. There is additional longer range research on true artificial red and white blood cells which could theoretically compose a blood substitute with higher fidelity to human blood.

Unfortunately, oxygen transport (the function that distinguishes real blood from other volume expanders) has been very difficult to reproduce. There are two basic approaches to constructing an oxygen therapeutic:

• perfluorocarbons (PFCs), a chemical compound which can carry and release oxygen. The specific PFC usually used is perfluorocarbon.
• hemoglobin derived from humans, animals, or artificially via recombinant technology

Perfluorochemicals will not mix with blood, therefore emulsions must be made by dispersing small drops of PFC in water. This liquid is then mixed with antibiotics, vitamins, nutrients and salts, producing a mixture that contains about 80 different components, and performs many of the vital functions of natural blood. PFC particles are about 40 times smaller than the diameter of a red blood cell (RBC). This small size can enable PFC particles to traverse capillaries through which no RBCs are flowing. In theory this can benefit damaged, blood-starved tissue, which conventional red cells cannot reach. PFC solutions can carry oxygen so well that mammals and humans can survive breathing liquid PFC solution, called liquid breathing.

Hemoglobin is the main component of red blood cells, comprising about 33% of the cell mass. Hemoglobin-based products are called HBOCs (Hemoglobin Based Oxygen Carriers). However pure hemoglobin separated from red cells cannot be used since it causes renal toxicity. It can be treated to avoid this, but it still has incorrect oxygen transport characteristics when separated from red cells. Various other steps are needed to form hemoglobin into a useful and safe oxygen therapeutic. These may include cross-linking, polymerization, and encapsulation. These are needed because the red cell is not a simple container for hemoglobin, but a complex entity with many biomolecular features.^[1]

The first approved was a perfluorocarbon-based product called Fluosol-DA-20, manufactured by Green Cross of Japan. It was approved by the Food and Drug Administration (FDA) in 1989. Because of limited success, complexity of use and side effects, it was withdrawn in 1994. However Fluosol-DA remains the only oxygen therapeutic ever fully approved by the FDA.

In 1990s because of the risk of undetected blood bank contamination from AIDS, hepatitis C and other emergent diseases such as Creutzfeldt-Jakob disease, there was additional motivation to pursue oxygen therapeutics. Significant progress was achieved, and a hemoglobin-based oxygen therapeutic called Hemopure was approved for Phase III trial (in elective orthopedic surgery) in the U.S., and more widely approved for human use in South Africa.

In December 2003 a new hemoglobin-based oxygen therapeutic, PolyHeme, began field tests in a Phase III trial on emergency patients (in trauma settings) in the U.S. PolyHeme is the 15th experiment to be approved by the Food and Drug Administration since 1996. Patient consent is not necessary under the special category created by the FDA for these experiments. In late 2005, an independent panel verified, after the fourth and final review of 500 trauma patients enrolled in this study by that date, that no statistical evidence of safety concerns had arisen so far in the study. This pivotal study is expected to conclude in mid-2006 with final enrollment of 720 patients. If successful, this trial could lead to Food and Drug Administration approval of PolyHeme for use for severely bleeding trauma victims as early as sometime in 2007.

The U. S. Military is one of the greatest proponents of oxygen therapeutics, mainly because of the vital need and benefits in a combat scenario. Since oxygen therapeutics are not yet widely available, the United States Army is experimenting with varieties of dried blood, which takes up less room, weigh less and can be used much longer than blood plasma. Water has to be added prior to use. These properties make it better for first aid during combat than whole blood or packed red cells.

Advantages

Oxygen therapeutics even if widely available would not eliminate the use of human blood, which performs various functions besides oxygen transport. However oxygen therapeutics have major advantages over human blood in various situations, especially trauma.

Blood substitutes are useful for the following reasons:

1. Donations are increasing by about 2-3% annually in the United States, but demand is climbing by between 6-8% as an aging population requires more operations that often involve blood transfusion.
2. Although the blood supply in the US is very safe, this is not the case for all regions of the world. Blood transfusion is the second largest source of new HIV infections in Nigeria. In certain regions of South Africa as much as 40% of the population has HIV/AIDS, and thorough testing is not financially feasible. A disease-free source of blood substitutes would be incredibly beneficial in these regions.
3. In battlefield scenarios it is often impossible to administer rapid blood transfusions. Medical care in the armed services would benefit from a safe, easy way to manage blood supply.
4. Great benefit could be derived from the rapid treatment of patients in trauma situations. Because these blood substitutes do not contain any of the antigens that determine blood type, they can be used across all types without immunologic reactions.
5. While it is true that receiving a unit of transfused blood in the US does not carry many risks, with only 10 to 20 deaths per million units, but blood substitutes could eventually improve on this. There is no practical way to test for prion transmitted diseases in donated blood, such as Mad Cow and Cruetzfeld-Jacob disease, and other disease could emerge as problems for the blood supply, including Smallpox and SARS.
6. Transfused blood is currently more cost effective, but there are reasons to believe this may change. For example the cost of blood substitutes may fall as manufacturing becomes refined.
7. Blood substitutes can be stored for much longer than transfused blood, and can be kept at room temperature. Most hemoglobin-based oxygen carriers in trials today carry a shelf life of between 1 and 3 years, compared to 42 days for donated blood, which needs to be kept refrigerated.
8. Blood substitutes allow for immediate full capacity oxygen transport, as opposed to transfused blood which can require about 24 hours to reach full oxygen transport capacity due to 2,3-diphosphoglycerate depletion.

Current oxygen therapeutics under development

Perfluorocarbon based

• Oxygent, by Alliance Pharmaceutical. Status: U.S. phase II trials, European phase III trials

Oxygent is a solution used as an intravascular oxygen carrier to temporarily augment oxygen delivery to tissues and is currently being developed by Alliance Pharmaceutical Corp. Right now, the goal of the development of Oxygent is simply to reduce the need for donor blood during surgery, but this product clearly has the potential for additional future uses. Perfluorocarbons surrounded by a surfactant called lecithin and suspended in a water based solution give Oxygent its oxygen carrying capacity. The Oxygent particles are removed from the bloodstream within 48 hours by the body's normal clearance procedure for particles in the blood. Namely, the lecithin is digested intracellularly and the PFC's are exhaled through the lungs. The fact that this blood substitute is completely man-made gives it certain distinct advantages over blood substitutes that rely on modified hemoglobin, such as unlimited manufacturing capabilities, ability to be heat-sterilized, and the PFCs’ efficient oxygen delivery. Oxygent has done well in most clinical trials, but recently ran into some trouble, with participants in a cardiac surgery study slightly more likely to suffer a stroke if treated with Oxygent rather than the standard care.

• Oxycyte, by Synthetic Blood International. Status: U.S. phase II trials
• PHER-02, by Sanguine Corp. Status: In research
• Perftoran (Russian). Status: approved for Russian clinical application in 1996

Hemoglobin based

• Hemopure, by Biopure Corp. Status: U.S. phase III trials, more widely approved in South Africa.

Hemopure is made by Biopure, one of the leading companies in the development and manufacture of oxygen carrying solutions. It is Biopure’s first-in-class product for human use, and is a Hemoglobin based oxygen carrying solution (HBOC). It is made of chemically stabilized, cross-linked bovine (cow) hemoglobin situated in a salt solution, and many safety measures are taken to ensure that the product is safe and free of pathogens, including herd control and monitoring. Hemopure molecules can be up to 1,000 times smaller than RBC’s facilitating oxygen transport and off-loading to the tissues. Hemopure is currently in Phase III clinical trials in the US, and is approved for use in South Africa for the use of surgical patients who are anemic, thereby reducing or eliminating the need for blood transfusions for these patients.

• Oxyglobin, by Biopure Corp., approved for veterinary use in US and Europe.

Manufactured by Biopure, Oxyglobin solution is the first and only oxygen therapeutic to be both US FDA and European Commission approved for veterinary use. The solution consists of chemically stabilized bovine hemoglobin in a balanced salt solution and contains no red blood cells. The cross-linked hemoglobin, several tetramers bound together, works by circulation in the plasma and supplying oxygen to tissues. Introduced to veterinary clinics and hospitals in March of 1998 and nationally distributed by October 1998, Oxyglobin has been used primarily for blood transfusions and for treatment of anemia in dogs. Currently, Oxyglobin can only be used in canines and not in humans. The current supply of Oxyglobin is low, because the company is spending most of its resources on Hemopure, a blood substitute designed for human use.

• PolyHeme, by Northfield Laboratories. Status: Awaiting top-line results for its U.S. phase III trials

Polyheme is a hemoglobin based oxygen carrier and, as the only blood substitute to have completed a Phase III trial, represents the leading technology in this field. Developed and manufactured by Chicago based Northfield Laboratories, Inc., Polyheme originally began as a military project following the Vietnam War and has since shown great potential for both military and civilian use. Polyheme utilizes human hemoglobin as the oxygen carrying molecule in solution, and the extraction and filtration of this hemoglobin from red blood cells is the first step in production. Then, using a multi-step polymerization process, the purified hemoglobin is associated into tetramers and, as the final step, is incorporated into an electrolyte solution. The polymerization of the hemoglobin represents the critical step in this process because, as demonstrated by failed attempts at blood substitutes, when hemoglobin remains disassociated, it tends to take up nitric oxide, causing vasoconstriction. Also, free hemoglobin can be taken up by the kidney causing liver dysfunction and failure.

Recently, Northfield Laboratories has come under scrutiny for a Phase III trial they are conducting in over 20 level 1 trauma centers across the country. The controversy arises from the fact that the participants in this study are incapable of giving their consent due to the nature of their injuries. Even though this practice is sanctioned by the FDA as necessary emergency research, patients’ rights groups have begun to protest the study.

• Hemospan, by Sangart, Status: US Phase II trials

Hemospan is produced by the company Sangart, which was founded by Dr. Robert M. Winslow in 1998. Produced in powder form, the powder can then be mixed into liquid form and transfused immediately, regardless of a patient’s blood type. This technology relies on coupling with polyethylene glycol (PEG) to eliminate the toxicity associated with free hemoglobin. Sangart believes their product can be stored for years and that they have optimized certain factors involved in oxygen delivery in the production of Hemospan, so that their product ultimately presents the right amount of oxygen to the blood vessel wall. In the past four years, Hemospan has shown promise as a possible commercial product, yielding positive results in both Phase Ib/II and Phase II clinical trails.

• Dextran-Hemoglobin, by Dextro-Sang Corp.

Created by the Dexto-Sang Corporation, Dextran-Hemoglobin is a conjugate of the polymer dextran with human hemoglobin molecules. The safety of dextran has already been established, due to its wide use as a plasma volume expander. Conjugation of hemoglobin to dextran increases its half-life inside the body, and prevents tissue damage that occurs with free hemoglobin from processing by the kidneys and exit into the extracellular space.

Dextran-Hemoglobin is currently undergoing trials in dogs in Thailand, and the company hopes to begin human trials by the end of the year.

Hemotech by [HemoBiotech]

HemoBiotech, based in Dallas, TX is developing HemoTech, a human blood substitute developed in 1985 by researchers, Mario Feola, MD and Jan Simoni, PHD, DVM from the Texas Tech University Sciences Center (TTUHSC0). HemoBiotech has been able to identify and nullify the source of toxicity issues associated with previous blood substitute candidates. HemoTech, which is expected to be compatible will all blood types and a shelf life of 180+ days compared to 42 days for donated human blood. HemoTech's lack of toxicity is believed to be due to Hemobiotech's proprietary chemical modification of hemoglobin. The company believes the use of bovine blood provides and additional advantage over products developed from outdated human red blood cells or from perfluorochemicals (PFC's), as bovine blood is more readily available and more cost-effective to use. Limited tests has shown to be clear of the vasoconstriction and inflammatory toxicity issues that have hampered competitors.

Promising techniques

• Stem Cells

Recently the scientific community has begun to look at the possibility of using stem cells as a means of producing an alternate source of transfusable blood. A study performed by Giarratana et al describes a large-scale ex-vivo production of mature human blood cells using hematopoietic stem cells, and may represent the first significant steps in this direction. Moreover, the blood cells produced in culture possess the same hemoglobin content and morphology as do native red blood cells. The authors of the study also contend that the red blood cells they produced have a near-normal lifespan, when compared to native red blood cells—an important characteristic that current hemoglobin-based blood substitute fall short on.

The major obstacle with this method of producing red blood cells is cost. At the moment, the complex three step method of producing the cells would make a unit of these red blood cells too expensive. However, the study is the first of its kind to demonstrate the possibility of producing red blood cells which closely resemble native red blood cells on a large scale.

• Dendrimers

Researchers at the Dendritech Corporation have begun research, aided by a 2 year, $750,000 grant from the US Army, into the use of dendrimers as substitute oxygen carriers. The precise nature of the research cannot be disclosed, as the company’s patent application has not yet been approved. Researchers hope that dendrimer technology will be the first truly cost-efficient blood substitute.

• Biodegradable Micelles

In order to enhance circulation times, recombinant or polymerized hemoglobin can be encapsulated within micellar forming amphiphilic block copolymers. These systems are typically between 30-100 nm in diameter. The hydrophobic core of the polymer micelle is able to solubilze the similarly hydrophobic hemoglobin protein, while the water soluble corona (which is usually poly (ethylene glycol)) provides a steric barrier to protein absorption and provides protection from clearance by the reticuloendothelial system (RES).

• Placental umbilical cord blood

Cord blood collected asceptically from the placenta after the birth of a healthy baby can be used safely as a blood substitute. It has a higher hemoglobin content and growth factors then normal blood from an adult, which has the potential to benefit patients in varying diseases.

Withdrawn oxygen therapeutics

• Flourasol-DA, by Green Cross. Status: withdrawn in 1994 due to usage complexity, limited clinical benefit and complications
• HemAssist, by Baxter International. Status: withdrawn in 1998 due to higher than expected mortality
• Hemolink, by Hemosol, Inc. Status: phase III clinical trials were discontinued in 2003 when cardiac surgery patients receiving the product experienced higher rates of adverse cardiovascular-related events. Some very limited ongoing investigation is still being conducted as of 2007, including the possibility of a future modified Hemolink product.

Clinical Trials of Blood Substitutes

Since blood transfusions are often most critically needed in trauma situations where obtaining informed consent is either difficult, impossible, or a barrier to providing lifesaving care, the ethics of clinical trials for these products are difficult. In the United States, these trials are performed under a specific exemption from requirements for informed consent ^[2].

Nominally, the community where blood substitutes will be used in clinical trials is informed, and wristbands or other clear markers are distributed so that a person can "opt out" of the trial. The concept that a person could be involved in experimentation on humans without consent is controversial^[3].

See also

• Blood transfusion
• Bloodless surgery

References