AU4264100A - Hemodilution facilitated by mounting oxygenation status - Google Patents

Description

AUSTRALIA

PATENTS ACT 1990 DIVISIONAL APPLICATION NAME OF APPLICANT: ALLIANCE PHARMACEUTICAL CORP.

ADDRESS FOR SERVICE: DAVIES COLLISON CAVE Patent Attorneys 1 Little Collins Street Melbourne, 3000.

INVENTION TITLE: HEMODILUTION FACILITATED BY MONITORING OXYGENATION STATUS The following statement is a full description of this invention, including the best method of performing it known to us: 1A HEMODILUTION FACILITATED BY MONITORING OXYGENATION STATUS FIELD OF THE INVENTION The present invention relates to improved formulations relating to hemodilution. The improved formulations include an oxygen carrier in amounts based on the continuous monitoring of the mixed venous partial pressure of oxygen (PvO 2 or other tissue oxygenation indices, and also include autologous blood or additional oxygen carrier to maintain the PvO 2 or other indices at or above a predetermined leveL BACKGROUND OF THE INVENTION More than 13 million units of blood are collected each year in the United States alone, and about million of these units are transfused into 4 million recipients. Of the transfused units, about two-thirds are used during surgical proceduresand the remainder are used primarily for treating severe anemia or in emergency indications. Experience from clinical studies suggests that postoperative recovery can be shortened if hemoglobin concentrations are not allowed to.fal to below 10 gidL the previously generally accepted indication for transfusion (Zauder, Anasth. Ci. North Amer. 8:471-80 (1990)). This criterion, however, is currently being reevaluated due in part to a recent increase in awareness of the risks associated with 15 allogeneic blood transfusion (NIH Consensus Conference JAMA 260:2700-2703 (1988)). This has also resulted in a renewed interest in the use of autologous blood transfusion techniques, in particular predonation and acute normovolemic hemodiution (ANH).

Although autologous blood transfusion reinfusion of the patient's own blood) was first employed over 170 years ago, it was not until the early 1970s that its use became more widespread because of 20 growing concerns about the transmission of hepatitis. More recently, interest in autologous transfusions on the part of both patients and physicians has been stimulated by the emergence of AIDS. Despite an increased awareness and acceptance of the benefits of autologous blood transfusion, recent studies have revealed the widespread underutization of autologous predonation (which is estimated to represent only of al units drawn nationwide).

S 25 ANH is a procedure whereby several units of blood are withdrawn from the patient at the beginning of surgery and simultaneously replaced with either a crystaloid or a coloid plasma volume expander (Stehing et aL Transfuhi 31:857 (1991)). The basic mechanism that compensates for most of the decreased oxygen capacity of the diluted blood is the rise in cardiac output and increased organ blood flow, factors that result from the improved fluidity of blood lower viscosity) at lower hematocrit levels (Mssmer et al Eir. Srg.

Rs. 18:254-263 (1986)). Weisskopf, Transfusin 35(1):3741 (1995) describes a mathematical analysis of acute isovoltmic hemodiution prior to surgical blood loss, which was used to detemine the magnitude of potential reductions in alogeneic transfusion. Weisskopf concluded that isovolomic hemodiution prior to surgery can obviate alogeneic blood transfusion or diminish the amount transfused.

Predonation typically involves withdrawal of several units of a patient's blood during the six weeks prior to surgery. To avoid excessive anemia, the amount of blood that can b safely predonated in the weeks before surgery is mited, as is the amount of blood that can be removed during ANH.

Quite apart from ANH and predonation, it has been suggested that red cell substitutes, or blood substitutes, could be used in place of allogeneic blood blood from other humans) during surgery.

Methods for facilitating autologous blood use which employ a synthetic oxygen carrier or blood substitute are disclosed in U.S. Patent No. 5,344,393 (Roth et al). Extensive research in the field of such blood substitutes over the past two decades has resulted in several candidate compositions. These include perfluorocarbon emulsions, such as FLUOSOL (Green Cross Corporation, Japan) and OXYGENT (Alliance Pharmaceutical Corp., San Diego, USA), and hemoglobin compositions, such as those derived from human, animal, or recombinant sources.

Traditional thinking has been that a red cel substitute would be given in volumes equal to the amount of whole blood that would be used for the same purpose. The use of such blood substitutes in large volumes to replace blood used in transfusions has not been entkely satisfactory in earlier applications. For example, early studies using FLUOSOL as a large volume blood substitute found that following blood loss, FLUOSOL was "unnecessary in moderate anemia and ineffective in severe anemia." Gould, et aL, New EngL J. Mad. 314:1653 (1986). In this study, the average increase in arterial oxygen content with the drug was 15 only 0.7 ml/deciliter. Thus, it was concluded that use of fluorocarbon emulsions as blood substitutes would not provide a significant benefit in severely anemic and bleeding patients. Indeed, although the U.S. Food S. Drug Administration approved FLUOSOL in 1989 for use as a perfusion agent to enhance myocardial oxygenation during percutaneous transluminal coronary angioplasty (PTCA), it did not approve an earlier application for use as a large volume blood substitute for general use.

20 The problem in using fluorocarbon emulsions and hemoglobin compositions as red cell substitutes or blood substitutes to compensate for blood loss from surgery, disease, or trauma lies in the relatively short circulating blood half life of those materials h wive. Healthy humans typically require about two weeks to manufacture new red cells and increase their bematocrit to normal levels following blood loss. In contrast, i* the intravascular half life of fluorocarbon emulsions and hemoglobin substitutes i vive is typically less than 25 72 hours, most often much less than 24 hours. Thus, even if sufficient quantities of a red cae substitute are administered during andlor after surgery, for example, to provide adequate oxygen delivery, the oxygen carrying capacity wll drop significantly long before the body can compensate by making new red cells. One aspect of the current invention therefore defines an improved method to use red cel substitutes or blood substitutes for temporary short-term perioperative use in conjunction with autologous blood conservation strategies as a means of reducing or eFninating allogeneic blood transfusions.

Treatment of intracoronary thrombotic events such as myocardial infarcts usualy involves systemic adnristration of thrombolytic agents, for example tissue plasminogen activator (tPA) or streptokinase.

Mechanical intervention using percutaneous coronary angioplasty (PTCA) is also used. Under no circumstance during current treatment methods is blood purposefully diuted, as this would diute the concentration of red blood cels and thus impair the delivery of oxygen to the heart Many clular elements of blood, however, are detrimental in the case of myocardial infarction. For example, it is wel known that platelets are necessary for the process of thrombus formation; reduction in the number of platelets would result in attenuation of the rate of thrombus formation following infarction. Further, certain white blood cells, polymorphonuclear leukocytes (neutrophils), are known to be activated at the site of the infarct to release cytotoxic components including oxygen free radicals, which, upon successful opening of the stenosed artery, are responsible for damaging normal cells through a phenomenon known as reperfusion injury. It would be beneficial, therefore, to dilute blood during and for a specified time after treatment of a myocardial infarction in order to reduce the number of platelets and neutrophils that exacerbate the effects of the infarct.

Hemodiution is not done, however, because it is also necessary to maintain high red blood cell levels to deliver oxygen to the myocardium.

The current invention therefore also defines an improved method to use red cel-substitutes or blood substitutes for temporary short-term use in conjunction with treatment of myocardial infarction as a means of reducing or eliminating the detrimental effects associated with the infarct while providing enhanced oxygen deivery to the tissues.

SUMMARY OF THE INVENTION 15 The present invention provides a composition for use in a method for facilitating autologous blood use by a patient facing a loss of blood, comprising the steps of: removing and storing a portion of the patient's blood while intravenously administering a biocompatible iquid in sufficient quantity to bring the patient's blood hemoglobin level to a desired concentration; intravenously administering a biocompatible oxygen carrier, while periodically or continuously assessing the patient's tissue oxygenation, after which the patient 20 undergoes a further loss of blood; and intravenously readministering the stored blood to the patient in response to the oxygenation measurements to maintain oxygenation measurements at or above a desired value. In one embodiment, the biocompatible liquid comprises a hemodluent. In another embodiment, the hemodiuent is administered separately from the oxygen carrier. The method further comprises the step of administering additional oxygen carrier in response to the oxygenation assessments to maintain oxygenation assessments at or above a desired value prior to readministering the stored blood. The oxygen carrier is preferably derived from human, animal plant, or recombinant hemoglobin, or it may be a fluorocarbon emulsion.

When the oxygen carrier is a fluorocarbon emulsion, the volume of the administered oxygen carrier is advantageously less than 50% of the volume of the biocompatible iquid. The fluorocarbon emulsion preferably has a concentration of at least 40%, preferably 50% or 60% wlv.

The biocompatible iquid is advantageously selected from the group consisting of a crystalloid, a coloid, a biocompatible oxygen carrier, and combinations thereof. The method also may further comprise the step of administering oxygen breathing gas to the patient during the procedure. The blood loss is often blood loss associated with surgery. Alternatively, the blood loss is associated with trauma.

The amount of oxygen carrier administered is usualy between about 0.5 and 10 mllkg, based on the body weight of the patient. The desired concentration of hemoglobin may advantageously be about 8 4gdL The assessing of the patient's tissue oxygenation can be performed by assessing PvO 2 such as by using a pulmonary artery catheter. Preferably, the desired value of PvO 2 referred to above is about mmHg.

The present invention also includes a composition for use in a method for the treatment of organ ischemia or infarct, including myocardial infarction, comprising the steps of removing a portion of the blood of a patient in need of treatment for organ ischemia or infarct and intravenously administering a biocompatible liquid in sufficient quantity to reduce the patient's blood hemoglobin level to a desired concentration; and intravenously administering a biocompatible non-red call oxygen carrier in conjunction with the removing step to maintain oxygenation of the patient's tissues at or above a predetermined level In one embodiment, the biocompatible liuid comprises a hemodiluent. In another embodiment, the hemodiluent is administered separately from the oxygen carrier. The oxygen carrier and biocompatible liquid may be the same or different, and may be as described above. The method advantageously also includes the step of administering oxygen breathing gas to the patient during the method. The amount of oxygen carrier ao administered is preferably between about 0.5 and 10 mUkg. based on the body weight of the patient. As 15 above, one preferred concentration of hemoglobin after hemodilution is about 8 gIdL In order to assure adequate oxygenation of tissues including myocardium, the method further comprises the step of assessing the patient's tissue oxygenation by assessing PvO 2 as discussed above, to maintain a desired value of PvO 2 at a value, for example, of about 40 mmHg. In one modification of the method, the oxygen carrier constitutes at least a part of the biocompatible liquid.

20 In addition to the foregoing, the invention comprises a composition for use in a method of hemodiluting a patient, comprising the steps of removing and storing a portion of the patient's blood while intravenously administering a biocompatible oxygen carrier and periodically or continuously assessing the patient's tissue oxygenation, after which the patient undergoes a further loss of blood, and administering additional oxygen carrier to the patient in response to the oxygenation assessments to maintain the oxygenation assessments at or above a desired value. The method may further comprise the step of readministering the stored blood to the patient The oxygen carrier and the desired values of oxygen carrier deivery and oxygenation may be as described above. The method may also include the step of administering oxygen breathing gas to the patient during the method.

Yet another aspect of the present invention comprises a composition for use in a method of hemodluting a patient, comprising the steps of removing and storing a portion of the patient's blood whie intravenously administering a biocompatible oxygen carrier and periodically or continuously assessing the patient's tissue oxygenation, after which the patient undergoes a further loss of blood. The method may further comprise the step of readministering the stored blood to said patient. The oxygen carrier and the desired values of oxygen carrier delvery and oxygenation may be as described above. The method may also include the step of administering oxygen breathing gas to the patient during the method.

BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 is a graph showing acceptable blood loss during surgery without hemodilution, administration of allogeneic blood, or administration of a synthetic oxygen carrier, assuming a normal hemoglobin (Hb) concentration of 14gidL in the patient at the time of surgery, and a concentration of 10 gIdL being required at the end of operation. The calculated permitted blood loss before transfusion is deemed necessary amounts to 1682 mL Figure 2 is a graph showing acceptable blood loss during surgery using conventional hemodiution methods. It is assumed that no allogeneic blood is to be given, that initial ANH is to a Hb of 10 gmdL Intraoperative transfusion of ANH blood occurs at a Hb of 8 gmidL and a Hb of 10 gm/dl is given at the end of operation. The calculated permitted blood loss amounts to 2366 mL Figure 3 is a graph showing acceptable blood loss during surgery using the method of isovolemic hemodilution described by Weisskopf, Transfusion 35(1h37-41 (1995). This method allows a blood loss of 2500 mL Figure 4 is a graph showing acceptable blood loss during surgery using the method of the present 15 invention, which allows a blood loss of 4000 mL The present example uses 1.8 gmlKg of a perflubron emulsion given at 8 gmldL hemoglobin concentration. This method assumes that initial ANH is to a Hb concentration of 8 gmldL As surgical blood loss starts, ANH blood is transfused to keep the Hb at 8 gmndL Figure 5 is a graph showing the relationship between the 02 delivery from hemoglobin in blood and cardiac output under normal conditions (hematocrit Total 02 utilization (or consumption; VO 2 i S 20 equal to the product of cardiac output times the arterial venous 02 content difference, and is indicated by the cross-hatched area. OxyHb dissociation curves were generated from data provided by the model developed by Winslow, Int. J. Cbn. Monitor Conp. 281-93 (1985).

O: DETAILED DESCRIPTION OF THE INVENTION A. Overview of the Invention The invention described below combines the use of limited intravascular half-ife oxygen carriers (blood substitutes) with hemodilution methods to increase alowable blood loss during surgery. Increasing the alowable blood loss decreases the need for autologous or alogeneic blood transfusion, thereby reducing or elninating the attendant risks and complications. The invention also provides for adjunctive treatment of organ ischemia or infarct, including myocardial infarction, using hemodilution and administration of intravascular oxygen carriers.

In one embodiment of the present invention, blood is removed from the patient prior to initiation of a surgical procedure, and the removed blood is stored for later readministration to the patient The removed blood is replaced with an asanguineous fluid, generally crystaloid andlor cooid-based solutions which may also be the oxygen carrier red cml substitute based on hemoglobin (Hb) or fluorocarbon, to maintain nonnovolemia, while bringing the red cell contained hemoglobin concentration down to a predetamnined leveL At this point, the oxygen carrier is administered if not already administered as the hemodluent during ANH.

I

-6- Additional blood is removed from the patient while monitoring the mixed venous partial pressure of oxygen (Pv02) or other indices of global or regional tissue oxygenation. Tissue oxygenation can be assessed by use of oxygen electrodes, NADH fluorescence, or other means. When the Pv02 or other index reaches a certain trigger level, surgery is initiated. During the surgical procedure Pv02 or other oxygenation indices are continuously or periodically monitored and the autologous blood is added back to the patient in response to the oxygenation level to maintain that level at or above the trigger leveL Alternatively, additional doses of the oxygen carrier can be administered until the maximum tolerated dose is reached.

The oxygen carrier is administered to the patient to supplement the oxygen-carrying capacity of the blood during or after hemodilution with crystalloid andlor colloid-based solutions, or the oxygen carrier can serve as the hemodiluent itself. In this clinical situation an idditional margin of safety is afforded to the hemodiluted patient, by augmenting total oxygen delivery.

The combined use of autologous and blood substitute infusion technologies to avoid allogeneic transfusion is emphasized. The present invention contemplates use of both predeposit and perioperative autologous technologies with preferably less than one-to-one volume infusions of various oxygen-carrying blood 15 substitute formulations. This invention includes use of any or al of these technologies in whatever order or of whatever magnitude they may be clinically useful in the perioperative clinical setting described.

S. Another aspect of the present invention provides compositions and uses thereof for the treatment of organ ischemia or infarct, including myocardial infarction. Both blood oxygenation and dilution are accomplshed for more beneficial adjunctive treatment This aspect of the invention involves the hemodilution 20 of the patient suffering organ ischemia with a generally crystalloid andlor coloid-based solutions. Blood is removed from the patient and replaced with an asanguineous fluid, while at the same time, the patient is administered an oxygen carrier red cel substitute, such as a fluorocarbon emulsion or hemoglobin solution As before, the crystaloid or colloid-based solution may also be the oxygen carrier red cel substitute based o. on hemoglobin (Hb) or fluorocarbon. The administration of the oxygen carrier ensures adequate delivery of oxygen to the heart and other tissues, while hemodilution reduces the number of platelets, neutrophils and other celular components that exacerbate the effects of the myocardial infarction. PvO 2 or other oxygenation indices are continuously or periodicaly monitored and additional doses of the oxygen carrier are administered unti the maximum tolerated dose is reached in response to the oxygenation level to maintain that level at or above the trigger level.

The oxygen carrier is administered to the patient to supplement the oxygen-carrying capacity of the blood during hemodiution with crystaloid andlor colloid-based solutions. In this cinical situation total oxygen delvery is augmented whle the number of detrimental cels in the blood is reduced.

One unique feature of the present invention is of particular importance. By monitoring the mixed venous partial oxygen pressure or other oxygenation indices during surgery or organ ischemia (rather than using conventional hemoglobin or hematocrit measurements), and by using a non-blood oxygen carrier, increased amounts of blood can safely be removed (below a conventional hematocrit-based transfusion trigger in the case of surgery). The present invention therefore increases the margin of safety of existing autologous transfusion technologies, by increasing the amounts of blood which can safely be lost during surgery, and more accurately determining the oxygenation status of the tissues. It also provides for augmenting oxygen delvery to the myocardium and other organs and tissues whie reducing the number of cells in the blood which exacerbate the damaging effects of the ischemia or infarct.

B. Materials A large number of materials suitable for use in the present invention are already known in the art.

Without imiting the scope of the invention, certain representative materials are discussed below.

Several compositions have been proposed or demonstrated to function as intravenous oxygen carriers. These include fluorocarbon emulsions, including but not finited to perfluorocarbon emulsions. Such emulsions are typically fluorocarbon-in-water emulsions having a discontinuous fluorocarbon phase and a continuous aqueous phase. The emulsions typically include emulsifying agents and osmotic agents, together with buffers and electrolytes.

The fluorocarbon emulsion may be selected from a wide range of suitable emulsions. Preferably, S 15 it is a fluorocarbon-in-water emulsion, having a preferred fluorocarbon concentration of about 5% to about 125% weight per volume (wlv) that is used.

Fluorocarbons are fluorine substituted hydrocarbons that have been used in medical applications as imaging agents and as blood substitutes. U.S. Patent No. 3,975,512 to Long discloses fluorocarbons, including brominated perfluorocarbons, used as a contrast enhancement medium in radiological imaging.

20 Brominated fluorocarbons and other fluorocarbons are known to be safe, biocompatible substances when appropriately used in medical applications.

It is additionaly known that oxygen, and gases in general are highly soluble in some fluorocarbons.

This characteristic has permitted investigators to develop emulsified fluorocarbons as blood substitutes. For a general discussion of the objectives of fluorocarbons as blood substitutes and a review of the efforts and problems in achieving these objectives see "Reassessment of Criteria for the Selection of Perfluorochemicals for Second-Generation Blood Substitutes: Analysis of StructurelProperty Relationship" by Jean 6. Riess, Ariifii Organs 8:34-56, (1984) The fluorocarbon, in one preferred embodiment, is a perfluorocarbon or substituted perfluorocarbon.

Fluorocarbon molecules used in these emulsions may have various structures, including straight or branched chain or cycic structures, as described in Riess, J, Artiicil Organs 8(1)44-56 (1984). These molecules may also have some degree of unsaturation, and may also contain bromine or hydrogen atoms, or they may be amine derivatives. The fluorocarbons may be present in the emulsion in any useful concentration, but usualy range from about 5% to 125% wlv. As used throughout, concentrations defined as weightivolume are understood to represent grmsm and weight per volume to represent gramsl00 mL Although concentrations as low as 5% wlv are contemplated, in a preferred embodiment the concentrations are at least 25% or 30%, preferably at least 40%. 50%, 55%, 60%, 75% or 80% wlv.

Emulsions of 60%, 85%, 90%, and 100% are particularly preferred. Preferred fluorocarbon emulsion f ormulations are those disclosed in U.S. Patent Nos. 4,865,836, 4,987,154, and 4,927,623, which are hereby incorporated by reference.

There are a number of fluorocarbons that are contemplated for use in the present invention. These fluorocarbons include bis(F-alkyl) ethanes such as C 4

F

9 CH -CH 4

CF

9 (sometimes designated OF-44E1, im

C

3 FqCH-CHC 6

F

13 and C 6

F

13

CH-CHC

6

F

13 ("F56E);cycfic fluorocarbons, such as CIOFIB decalin. "perfluorodecafln' or F-adamantane F-methyladamantane (OFMAO), F-i .3dimethyladamantane (FDMA"), FVdi-or F-trimethylbicyclo[3,3,1 Infnane ("nonanew); perfluorinated amninas. such as F-tripropylamineOFTPA') and F-tri-butylamine ('FTBA), F.4-methyloctahydroquinohzlfle (OFMOfl"), F-nmethyl~dcahydroioquinollne (FIUa), F-n-rnethyldecahydroquinoiIne ("FHO) F-n-cyclohexypuflolidine ('FCHP) and F-2.butyotetrohydrofuran ("FC.75"or 'RM1 flu'). Oth~er suitable fluaorcarbonsmay be selected from brominated perfluorocarbons. such as 1 .bromo-heptadecaf luoro-octane

(C

8

F

17 Br, sometimies designated perfluorooctylbromide, "PFOB", or 'perflubron"), 1 .bromopenta-decafluoroheptone

(C

7

F

15 BrL end 1bromotridecafluorohaxane(C6Fl3Br, sometimes known as perfluorohexylbromideor "PFHB"). Other brominated fluorocarbons are disclosed inUS Patent No. 3,975,512 to Long. Also contemplated are fluorocarbons having :nonf luorine substituents, such as perfluorooctyl chloride, perf luorooctyl hydride, and similar compounds having different numbers of carbon atoms, eg., 5.12 carbon atoms.

Additional fluorocarbons contemplated in accordance with this invention include perfluoroalkylated ethers or polyathars, such as (CFh)CFO(CF 2

CF

2 2

OCF(CF

3 2

(CF

3 2

CFD-(CF

2

CF

2 3

OCF(CF

3 0C 3 0CF(0 2

(C

3 2

CFO(C

2

CF

2

(C

6

F

1 h)0- Further. fluorocarbon-hydrocarbon compounds, such as, for example compounds having the general formula C 0 2 CnF2n, 1'~r~l CnF~i+ or FnF, ,F-CC where n and n' are the same or different and are from about 1 to about 10 (so long as the compound or a mixture containing the compound is a liquid at room temperature). Such compounds, for example, include C 8

F

17

C

2 11 5 and C 6

F

13 CH-CI1CBH 13 It wil be appreciated that esters, thioethers, end other variously modified mixed fluorocarbon-hydrocarbon compounds are also encompassed within the broad definition of "fluorocarbon' materials suitable for use in the present inve~ntion. Mixtures of fluorocarbons are also contemplated. Additional "fluorocarbons" not listed here, but having those properties described in this disclosure that would lend themselves to use in vivo in accordance with the present invention are also contemplated.

Emulsfying agents used in the emulsions of this invention may be anionic, cationic; or non-ionic surf actants or combinations thereof as are well known to those in the chemical arts, or the may be mixtures of synthetic compounds such as Pluronic F-68, a condensate of ethylene oxide with propylene glycol, as used in U.S. Patent No. 4,073,879 to Long. Fluorosurfectants, such as those described by J. Riess et alot? SyMosaim on Blood Substitutes, Montreal (May, 1987), are particularly suitable and can also be used. Emulsifyig agents may also be mixtures of the above agents. Particularly suitable emulsifiers may include natural amphipathic; compounds such as phosphoilpids, particularly phosphatidyicholne, wherein combined hydrophilic and hydrophobic properties enable the molecule to interface with both aqueous and fluorocarbon systems, thereby forming the emulsion droplets. There are various species of each class of phospholipids, such as the phospholipid cholines, comprising various pairings of saturated and unsaturated fatty acids in the glycerol structures. Phosphatidylcholine is an abundant natural material (lecithin) which may be purified from egg yolk, or may be produced synthetically (Avanti Polar Lipids, Pelham, AL).

Phospholipid emulsifiers, particularly egg yolk phospholipid and lecithin, are particularly preferred.

The phospholipid emulsifying agent is typically included in the range of from 2 to 14% w/v, usually increasing the phospholipid concentration with increasing fluorocarbon concentration. The preferred amount for an emulsion comprising 75% wlv bromofluorocarbon is 2.5 to 5% wlv and 3.5 to 10% w/v of phospholipid for an emulsion with 100% wlv bromofluorocarbon. In a preferred embodiment the phospholpid comprises at least 2% wlv of the emulsion.

Emulsification requires large amounts of energy to convert a two-phase immiscible system into a suspension of discontinuous smal droplets of hydrophobic fluid in an aqueous continuous phase. Fluorocarbon emulsification may be carried out generaly by either of two general processes which provide energy to the 15 system to break up the fluorocarbon volume into small droplets. In sonication emulsification, a probe is inserted into the mixture of fluorocarbon, emulsifier, and aqueous phase, and bursts of energy are released from the ti of the probe. In a mechanical emulsification process, such as that performed by a MICROFLUIDIZER apparatus (Microfluidics, Newton, MA 02164), streams of the mixed emulsion components are directed through the apparatus at high velocity and under high pressure 15,000 psi), and the high shear forces or cavitation resulting from the mechanical stress applied to the fluid produce the emulsion.

SThe aqueous phase of the emulsion may have components dissolved therein which give the emulsion S..desirable properties. For example, it may comprise an osmotic agent to bring the emulsion to physiological isotonicity. The osmotic agent may be sodium chloride, or it may be a polyhydroxyl compound, such as a sugar or mannitoL The aqueous phase will also contain soluble buffering agents.

The ipid phase of the emulsion may also have components dissolved therein. For example, a phosphatidyl choline emulsifier may have glycerol phosphatidyl glycerol, other phosphoipids or cholesterol admixed, and further contain an antioxidant substance, such as a tocopherol, to protect against ipid oxidation.

Several fluorocarbon emulsions have been produced commercially for use as intravascular oxygen carriers. These include a mixed decaih emulsion formerly sold by Alpha Therapeutics Corp, Los Angeles, Calfornia under the trademark FLUOSOL and perflubron-based emulsions produced by ABance Pharmaceutical Corp. of San Diego, Calfomia, under the trademark OXYGENT.

One exemplary perflubron emulsion is a 90% (wiv) perflubron emulsion (Aliance Pharmaceutical Corp., San Diego, CA) having the folwing Formula I: FORMULA I PERFLUBRON EMULSION Component Percent (wlv) Perflubron 90.000 Egg Yolk Phospholipid 4.000 NaH 2 P0 4

*H

2 0, USP 0.052 Na 2 HPO427H 2 0, USP 0.355 NaCI, USP 0.280 EDTA, USP 0.020 d-a-tocopherol, USP 0.002 Water for injection 48.400 Hemoglobin compositions contemplated for use in the present invention are wae known. Such compositions are disclosed, for example, in the following U.S. Patents, which are hereby incorporated by reference: U.S. Patent Nos. 4,911,929; 4,861,867; 4,857,636; 4,777,244; 4,698,387; 4,600,531; 4,526,715; 4,473,494; and 4,301,144.

15 Various materials have been used successfully as plasma expanders in connection with hemodilution procedures. These include the wel-known categories of crystabid compositions (exempified by Ringerslactate and saline both from Baxter Healthcare Corp, Deerfield, IL) and colloid compositions. Colloid compositions include modified fluid gelatins, such as those sold under the following trademarks: PLASMAGEL Belon Lab., Neuily-sur Seine, France), GEUFUNDOL (Biotest, Frankfurt, Germany), 20 GELOFUSINE (Braun, Melsungen, Germany) and HAEMACEL (Hoechst-Roussel Pharmaceutical Inc., Sommervie, NJ); dextran solutions, such as those sold under the trademarks MACRODEX (dextran-70) and RHEOMACRODEX (dextran40) both from Pharmacia, Piscataway, NJ; albumin solutions, such as those sold under the trademark ALBUTEIN (Alpha Therapeutics, Los Angeles, CA) and human serum albumin S: 25 from Abbott Labs, North Chicago, IL; starch solutions such as Hetastarch (Hydroxyethylstarch), HAES 25 (Fresenius, Hamburg, Germany) and HESPAN (DuPont, Wilmington, DE). These are administered in various volumes to maintain the patient's blood volume in the normal range and to encourage the increase in cardiac output that accompanies hemodilution procedures. In general, crystaoid-based solutions need to be given in volume ratios of 2:1 or 3:1 to blood withdrawn; colloids are usualy given in lesser amounts.

C. Procedures Autologous blood use virtually eliminates the poisiity of contracting blood-borne diseases associated with transfusions as wed as transfusion reactions occurring as a result of incompatiblity between donor and recipient blood. Autologous blood for use in subsequent transfusions can be obtained in a number of ways, including one or more of the following: predeposit; perioperative isovolemic hemodilution; and intraoperative salvage.

Predeposit requires that the surgery be planned wel in advance of the actual date. Blood is donated by the patient during the weeks before surgery, and is stored for subsequent administration to the patient. Phlebotomies of 350400 ml are typically performed at 2-7 day intervals, with the last collection more than 72 hours before surgery. The blood may be stored in the liquid state as whole blood, or it may be divided into red cells and plasma which can be frozen to preserve lable components.

Perioperative isovolemic hemodution is the process of collecting blood immediately before a surgical procedure with the concomitant replacement by a sufficient volume of crystalloid or colloid solution. This practice decreases blood viscosity during surgery, thereby reducing the work load on the heart allowing cardiac output to rise and improving microcirculatory oxygen flow and distribution. Typically,.sufficient blood is removed to reduce the hemoglobin concentration from a typical normal value of approximately 14 g/dL to about 10 gIdL This blood is stored for readministration to the patient during or after surgery. After removal of some of the blood, or simultaneously with the removal a crystaloid or colloid plasma expander (or both) is administered to the patient to maintain blood volume at a desired value, typically at the normal value Intraoperative blood salvage involves collecting blood lost from a wound or body cavity during surgery, processing it, and reinfusing the processed blood into the same patient. This procedure is safe and effective if certain basic precautions are folowed to ensure against contamination of the blood with bacteria S 15 or other pathogens, or malignant cells. Autotransfusion devices for collecting, filtering, and reinfusing the blood are commercially available. Also, some devices separate and wash the red blood cells, thereby avoiding administration of blood contaminated by debris, irigating solutions, activated factors, anticoagulants, and free hemoglobin. Suitable devices of this type are exemplified by the Haemonetics Ceo Separator and Cel Washer, e Haemonetics Corp., Braintree, MA.

20 Detailed reviews of autologous blood procedures and acute isovolemic or normovolemic hemodilution are found, for example, in Stehling, et al, Transfusn 31:857 (1991) and Mercurial, et al Autoogous Blood, Transmedica Europe Limited, Eastbourne, United Kingdom (1991), which are hereby incorporated by reference.

In the practice of the present invention, autologous blood procedures (preferably involving perioperative hemodlution) are combined with administration of non-blood oxygen carriers, including hemoglobin compositions and, more preferably, fluorocarbon emulsions, together with the monitoring of the partial oxygen pressure in the venous blood (PvO 2 or other oxygenation indices in the patient.

Though it is generally accepted that venous blood oxygen tension reflects, but does not measure,

PO

2 of the tissue from which it is issuing, it is generally impractical except under unusual circumstances, to monitor PO 2 in venous blood draining from individual tissues or organs. Hence, the mixed venous PO 2 (PvO 2 is usually taken as an acceptable estimator of the oxygen deiverylconsumption ratio in the whole body and is used as a guide to the oxygenation status of the whole body. It would be logical therefore to use Pv0 2 as an indication for the need for blood transfusion during surgical procedures and in the trauma ituation.

During the perioperative period, blood transfusions are routinely administered when e "critical" hemoglobin (Hb) concentration or hematocrit is reached. This level has traditionally been at a Hb concentration of 10 gidL To determine the lowest acceptable Hb level and the level of a suitable transfusion trigger, it is necessary to first consider the changes that take place during hemodilution as blood is removed and normovolemia is maintained.

As a patient is hemodiluted, either intentionally as part of an autologous blood conservation program, or folowing surgical bleeding with maintenance of normovolemia, both Hb concentration and arterial 02 content (Ca02) decrease. As the red cell concentration falls, a reduction in whole blood viscosity occurs; this, together with the simultaneously occurring increase in venous return, causes a rise in cardiac output (CO) and an improvement in total 02 transport to the tissues (P02). The degree to which this physiological compensation occurs will primarily depend on the response of CO to the reduction in red cell mass. Some authorities have concluded that the relationship between decrease in Hb concentration and CO is linear whereas others have maintained that it follows a curvilinear relationship; the degree of curvature found is very minimal, causing many researchers to perform calculations that assume a linear relationship.

In man, the extent to which cardiac output increases as Hb concentration decreases varies between 0.25 iters per minute per gm of Hb change to 0.70 Uminig. Hence, the cardiac output response to hemodilution differs between patients and this will affect the Hb level at which additional oxygen carrying 15 capacity in the blood wil be needed. The necessity for transfusion of red blood cells will also vary depending on such factors as vascular tone, which will cause the viscosity contribution to total systemic resistance to vary, and the ability of the myocardium to function at low Hb levels. During moderate hemodiution, myocardial blood flow increases proportionately more than total cardiac output and hence, in the absence of significant coronary atherosclerosis, no myocardial ischemia occurs. It has been shown, however, that 20 low postoperative hematocrit (Hct) may be associated with postoperative myocardial ischemia in patients with generalized atherosclerosis. Though attempts have been made to define a critical Hct level an empiric automatic transfusion trigger should be avoided and red cel transfusions should be tailored to the individual patient and be triggered by his or her own response to anemia.

As arterial blood passes through the tissues, a partial pressure gradient exists between the P02 of 25 the blood in the arteriole entering the tissue and the tissue itself. Oxygen is, therefore, released from hemoglobin in the red cells and also from solution in the plasma; the 02 then diffuses into the tissue. The P02 of the blood issuing from the venous end of the capilary cylinder wl be a reflection of, but not necessarily equal to, the PO 2 at the distal (venous) end of the tissue through which the capillary passes.

Under normal conditions this is essentially the same as that of interstitial fluid in contact with the outside of the capilary. The degree of equilibration between blood and tissue may depend on the speed of passage of blood through the capilary bed and under conditions of critical oxygen deivery caused by extreme anenma, there may not be time for equilibration of tissue and blood PO 2 levels; this may lead to higher than expected mixed venous PO 2 (P0 2 Nevertheless, in the clinical situation, it is generally accepted that probably the most relable single physiological indicator for assessing the overall balance between oxygen supply and demand is mixed venous oxygen tension. It is therefore sensible to use PvO2 as an indication of the overal adequacy of tissue oxygenation and to use it as a transfusion trigger rather than to use the traditional "10130 rule" as an indication for red blood cell transfusion.

If PvO 2 is accepted as a reasonable indicator of patient safety, the question arises as to what can be considered a "safe" level of this parameter. Though much data exists on critical oxygen delivery levels in animals, there is little to indicate what a critical PvO 2 might be in the clinical situation. The available data indicates that the level is extremely variable. For instance, in patients about to undergo cardiopuknonary bypass, critical Pv02 varied between about 30 mm Hg and 45 mm Hg; the latter value is well within the range of values found in normal, fit patients. Furthermore, shunting of blood in the tissues will cause elevated levels of PvO 2 such as is found in patients in septic shock, and will result in 02 supply dependency.

A Pv0 2 value of 35 mm Hg or more may be considered to indicate that overall tissue oxygen is adequate, but this is implicit on the assumption of an intact and functioning vasomotor system. This PvO 2 level is reached at a Hb of about 4gidL in patients with good cardiopumnonary function; even lower PvO 2 levels are tolerated in some patients when increased fractional inspired 02 concentrations (Fi0 2 S) are employed. In each situation it is necessary to maintain a good margin of safety and it is best to pick a :O Pv02 transfusion trigger at which the patient is obviously in good condition as far as oxygen dynamics are o*oo 15 concerned.

Physiological and clinical studies involving measurement and calculation of oxygenation parameters are usually carried out using cardiac output measurements obtained by thermodilution using a pulmonary artery catheter such as a Swan-Ganz catheter. Oxygen delivery and oxygen consumption (VO 2 are then derived from measured or calculated arterial and mixed venous oxygen contents by using the Fick equation.

The Fick equation alows the determination of oxygen consumption based on the difference between arterial and venous oxygen content times cardiac output The equation is as folows: V0 2 (C0 2 Cv2) x CO where V0 2 oxygen consumption, CaO2 arterial oxygen content. C0 2 venous oxygen content, and CO cardiac output.

Accordingly, one embodiment of the present invention involves removal of a portion of the patient's blood, and administration of an intravenous fluid to reduce the patient's hemoglobin concentration from about the normal level of about 14 gIdL to a first "trigger point The intravenous fluid preferably includes a plasma expander, such as a coloid or crystaloid solution which may also be the oxygencarrier red cel substitute or blood substitute based on Hb or PFC. This blood removal is usualy delberate, although the invention may also be used with trauma victims or other patients suffering involuntary blood loss. Wth deliberate removal the blood is stored for readministration to the patient at a later time.

When the hemoglobin level reaches the first "trigger" point, an oxygen carrier is administered intravenously if not already done as part of the ANH procedure. Additional blood is then removed, and PvO 2 andlor other indicators of tissue oxygenation is continuously or periodicaly monitored, for example by using a pulmonary artery catheter, unti the oxygenation reaches a second trigger point At that time, autologous blood can be administered to the patient to maintain oxygenation at or above the second trigger point, or additional doses of the oxygen carrier can be given until the maximum tolerated dose is reached. In some instances, the patient wil not reach the second trigger point as the initial dose of oxygen carrier is sufficient to maintain oxygenation above the second trigger point, and no additional oxygen carrier or autologous blood need be administered.

The oxygen carrier used is one other than red blood cells, preferably a biocompatible fluorocarbon emulsion of the type previously discussed, although hemoglobin compositions are also contemplated, as are other oxygen carriers.

Another aspect of the present invention provides for the use of a combination of hemodilution and administration of oxygen carrier as adunctive treatment of organ ischemia or infarct, including myocardial infarction. Frequently, higher concentrations of inspired oxygen are given to a patient who has suffered a myocardial infarct to assure maximum saturation of hemoglobin in red blood cels and thereby maximum delivery of oxygen to damaged and potentially damageable myocardial tissue. Under no circumstance, however, is the blood purposefully diluted, as this would dilute the concentration of red blood cells and the ability of the blood to carry oxygen to the heart. This is so even though it is known that other cellular elements of the blood are detrimental contributing to the damage caused by the myocardial infarct.

Platelets, for example, are necessary for the process of thrombus formation. Neutrophils are known to be activated at the site of the infarct to release cytotoxic components, including free radicals, which are responsible for damaging normal cells.

Accordingly, it would be beneficial to dilute blood during and for a specified period of time after treatment of a myocardial infarct in order to reduce the number of platelets and neutrophils that exacerbate the effects of the myocardial infarct, provided that adequate oxygen delivery to the myocardium and other tissues can be maintained.

The present invention provides for the hemodilution of a patient suffering from organ ischemia or infarct using a crystalloid- or coloid- based hemodiluent and intravenously administering a non-blood oxygen 25 carrier such as a hemoglobin composition or a fluorocarbon emulsion. Alternatively, the hemodiluent may be i the oxygen carrier. During hemodilution and the administration of the oxygen carrier, the patient's PvO 2 or other oxygenation indices is monitored, and the oxygen carrier is administered to maintain the PvO 2 or other oxygenation indices at or above a predetermined leveL This embodiment of the present invention involves removal of a portion of the patient's blood during andlor for a specified time during treatment of organ ischemia or infarct, and administration of an intravenous fluid to reduce the patient's hemoglobin concentration from about the normal level of about 14 gldL to a first "trigger" point The intravenous fluid preferably includes a plasma expander, such as a coloid or crystaloid solution which may also be the oxygen-carrier red cell substitute or blood substitute based on Hb or PFC.

The blood is stored for optional readministration to the patient at a later time. In one embodiment, where the intravenous fluid contains an oxygen carrier, no further hemodiution is done end the hemodiution procedure of the present invention is complete. This procedure reduces the quantity of circulating platelets and neutrophils, decreases the viscosity of the blood, and assures adequate perfusion of the tissues due to the added presence of the oxygen carrier.

In an optional embodiment of the organ ischemia or infarct treatment of the present invention, when the hemoglobin level reaches the first "trigger" point, an oxygen carrier is administered if not already done as part of the ANH procedure. Additional blood is then removed, and PvO 2 andlor other indicators of tissue oxygenation is continuously or periodically monitored, for example by using a pulmonary artery catheter, unti the oxygenation reaches a second trigger point. At that time, additional doses of the oxygen carrier can be given unti the maximum tolerated dose is reached to maintain oxygenation at or above the second trigger point, or the autologous blood can be administered to the patient.

In either hemodilution associated with surgery or hemodlution associated with the treatment of organ ischemia or infarct, the volume of intravenous fluid administered to the patient is at least about equal to 75%, preferably at least about 100% of the volume of blood removed from the patient More preferably, the volume of intravenous fluid is between about 150% and 300% of the volume of blood removed, depending on whether the fluid is predominantly a colloid or a crystalloid and depending on whether it consists of or contains the oxygen carrier. Alternatively, the volume of intravenous fluid administered to the patient is adequate to reduce the hemoglobin concentration of the patient to the trigger levels discussed above.

In one embodiment of the invention, the intravenous fluid comprises a major portion of a plasma expander and a minor portion of oxygen carrier. The volume ratio of administered expander to an oxygen S 20 carrier will range from 0:1 to at least 10:1, depending on whether the fluid is a crystaloid or a coloid, and on the composition of the oxygen carrier, the concentration of the oxygen carrier, P0 2 and cardiac output.

These ranges are most desirable when using a high concentration fluorocarbon emulsion, having at last about preferably at least about 50% or 60% fluorocarbon, wlv.

In one preferred embodiment, where a fluorocarbon emulsion such as perflubron-based emulsion is 25 used as the oxygen carrier, the total amount of actual perfluorocarbon administered to the patient is advantageously from about 0.5 glkg to about 10 gkg, preferably 1.6 glkg, based on the weight of the Spatient. When a 90% wlv or 100% wlv fluorocarbon emulsion is used, the volune of emulsion necessary to delver the desired dosage is about 0.25 or 0.255 mUkg to about 10 or 11 nmlkg, preferably about 1 to 6 mlkg. Simple calculation provides the preferred volume of emulsion when different concentrations of fluorocarbon are used.

The hemodiuted patient is preferably administered a breathing gas enriched in oxygen, preferably at least 50-60%, and most preferably 75% or 100% oxygen. The effects of the enriched breathing gas, increased cardiac output due to hemodiution, the oxygen carrier, and the dissolved oxygen in the aqueous phase of the circulating intravascular fluid and plasma al combine to supply enhanced levels of oxygen to the patient. The colective contributions of these factors to oxygen delvery in the patint are discussed in more detal in section 0. below.

During or after the surgical procedure or other condition resulting in blood loss, or following treatment of organ ischemia or infarct, the autologous blood removed from the patient (or the red cell portion thereof) can be readministered to the patient to maintain PvO 2 andlor other indices of oxygenation at or above the second trigger point. The oxygen carrier, meanwhile, is cleared from the circulation in a relatively short time, and its oxygen-carrying function is supplanted by the autologous transfusion of red cells, if required.

Accordingly, there are various trigger points that are important to the use of the present invention.

One is the hemoglobin or PvO 2 value at which oxygen carrier is infused if not already administered during the ANH. Others are the Pv02 values at which additional doses of the oxygen carrier or transfusion with autologous blood are initiated. Appropriate values in any particular instance or for any particular type of procedure will be determined with consideration of such variables as age, sex, weight, cardiac status, disease state, and so forth. In general however, one would expect that the first trigger point would occur during hemodilution at a hemoglobin level of between about 7 and 10 gidL typically at about 8 gIdL (Alternatively, it could occur at a PvO 2 value of about 35 mm Hg to about 45 mm Hg, preferably at about 40 mm Hg).

One would expect that the second trigger point would occur at a PvO 2 value of about 30 mm Hg to about 50 mm Hg, preferably at a value of about 40 mm Hg.

A comparison of the acceptable blood loss levels using conventional methods and using the present invention is shown in Figures 1-4.

Figure 1 is a graph showing acceptable blood loss during surgery without hemodilution, 20 administration of allogeneic blood, or administration of a synthetic oxygen carrier, assuming a normal hemoglobin concentration of 14gidL in the patient at the time of surgery, and a concentration of 10 gidL being required at the end of surgery. The hemoglobin concentration is generally not allowed to fal postoperatively below about 10 gIdL This allows a blood loss of 1682 mL before transfusion is deemed necessary.

25 Figure 2 is a graph showing acceptable blood loss during surgery using conventional hemodilution methods, wherein the hemoglobin concentration is alowed to fal to a level of about 8 gmldL This method allows for blood loss up to about 2366 mL Figure 3 is a graph showing acceptable blood loss during surgery using the mathematical analysis described by Weisskopf, Transfuson 35(1)3741 (1995). Assuming that hemodilution is completed before surgical blood loss is begun and that transfusion of removed blood is begun when surgical blood loss begins and lost blood is replaced at a rate that maintains the target hematocrit, this method allows for blood loss of 2500 mL Figure 4 is a graph showing acceptable blood loss during surgery using the present invention. By monitoring PvO 2 levels or other indices of tissue oxygenation and using them as an indicator of the overal oxygenation status of the patient, rather than hemoglobin or hematocrit measurements, and by administering an oxygen carrier, blood loss can safely be increased to 4000 mL The present example uses 1.8 gmlKg of a perflubron emulsion given at 8 gmldL hemoglobin concentration. This method assumes that initial ANH is to a Hb concentration of 8 gmldL As surgical blood loss starts, ANH blood is transfused to keep the Hb at 8 gmldL D. Oxygen Delivery to Tissues Although not intending to be bound by any particular theory of operation, the following discussion provides a framework for understanding the physical and physiological mechanisms contributing to the function of the present invention.

Oxygen transport to tissues can be considered to occur via two processes. The first is the convective (bulk) delivery of oxygen to tissues; the second is the delivery of oxygen to tissues via a diffusive process.

Convective Oxygen Delivery The first process, convective 02 delivery, is described by the Fick equation: V0 2 (CaO2 C,0 2 x CO Although the Fick equation is quite straightforward, a number of physiological variables of importance are imbedded in it. For example, the arterial.venous differential in oxygen content CaO2 Cv02 is determined by the 02 content of both arterial (Ca02) and venous (CvO 2 blood, respectively, which, in turn, are directly related to the hemoglobin (Hb) concentration and the 02 saturation and the contact of 02 in the plasma. Oxygen saturation is determined by the P02 and by the position of the oxyHb (oxygenated form of Hb) dissociation curve. The PO 2 is determined by the 02 tension in the inspired air and the capacity of 20 the lung to oxygenate pumonary caplary blood. Finaly, the position of the oxyHb dissociation curve is determined by 23-diphosphoglycarate (2,3-DPG) as wel as pH and pCO 2 which differ between arterial and venous blood and the temperature.

Similarly, cardiac output (CO) is controled by many factors, including heart rate, the left ventricular filing volume and ejection fraction stroke volume), and the demand for 02 in tissues oxygen 25 consumption, V0 2 Assuming a constant blood volume and under stable hemodynamic conditions, the left ventricular fiing volume is proportional to the blood viscosity, which, in normal humans, is priarily a i function of the hematocrit (percent of red ceb in blood).

Some of these complex relationships can be shown graphicaly (see Figure In Figure 5, 02 content is plotted against 02 tension, PO 2 Figure 5 presents data for a normal 70 kg man at rest with a hemoglobin concentration of 14.4 gIdl (hematocrit The data for the oxyHb dissociation curve used to create this graphic representation were generated by the model developed by Winslow (1985, which calculates the total 02 contents dissolved in the plasma and bound to hemoglobin. For a given arterial and venous PO 2 of 100 and 40 ton, respectively, the arterial to venous oxygen content difference (CaO 2 CvO 2 is 5 mUdL At a normal cardiac output of 5 Umin, the 02 consumption (VO2, represented by the crosshatched area) is approxiately 250 mUmin or 5 mUkglmin.

Normaly, more 02 is delvered to tissue than is utilized providing a "margin of safety." When the -18convective (bulk) delivery of 02 decreases below a certain critical point, tissue function may be compromised, with various consequences such as tissue hypoxia, production of lactic acid, infarction, necrosis, etc. Once this critical oxygen delivery level is reached when 02 delivery is severely limited), then V0 2 (oxygen consumption) will be supply.limited. The actual value for the critical oxygen delivery level is very difficult to specify, since there are likely to be different values for different organs or different capillary beds.

When 02 consumption is not supply-limited, changes in 02 content of the arterial blood can be compensated for by other normal physiological mechanisms. For example, in anemia, the cardiac output becomes elevated (see below), as does the level of red cell 23-0PG. The latter serves to shift the oxyHb dissociation curve to the right (reduced affinity, increased P 50 (the P02 at which hemoglobin is saturated with 02).

A similar compensatory mechanism (with respect to the cardiac output) occurs during acute normovolemic hemodilution (Messmer et aL Res. Exp. Med. 159:152-56 (1986)). As the hematocrit decreases during the hemodilution, blood viscosity also decreases significantly, which allows the cardiac output to increase without any significant changes in the work load on the heart. In this way, total oxygen transport (002) can be maintained.

Work by Guyton et aL (Cardac Output and its Regulatin, 2nd Ed. Saunders, Philadelphia (1973)) has shown that over a broad range, the cardiac output varies inversely with hematocrit. A hematocrit within the range of approximately 40 to 45% for normal resting humans is considered most appropriate. When hematocrit values exceed 45%, blood viscosity limits cardiac output such that there is lttle beneficial effect 20 from the additional 02 carrying capacity of the increased number of circulating red cells. When the hematocrit is less than about 40%, the lower viscosity results in a decreased total peripheral resistance to blood flow which allows cardiac output to increase in order to maintain normal oxygen delivery.

It should be noted that augmenting 02 transport by administration of a cel-free oxygen carrier differs from simple transfusion in several important ways. A key point in understanding the value of a low- S 25 dose acellular "blood substitute" is that olasma 0 2 is increased, rather than red cll 02, as is the case with transfusion of blood. Transfusion of red cells will increase bulk blood viscosity, which can cause a decrease in cardiac output and therefore may not increase the bulk 02 delivery.

Addition of a cell-free 02 carrier, on the other hand, wil increase bulk 02 deivery by elevating the 02 content of the plasma and potentially increasing the cardiac output (since overall blood viscosity would be reduced). This additional contribution to 00 2 is primarily due to an increased amount of 02 dissolved in the plasma compartment. 002 can be further increased by addition of a dose of perflbron emulsion or other oxygen carrier under these conditions which would provide an even greater margin of safety.

As a result, the hematocrit and hemoglobin levels can be significantly decreased when compared with the prior art methods, since the hemoglobin and hematocrit measurements do not adequately reflect oxygen carried in the added liuid volume and carried by the oxygen carrier. Nor do they account for increased cardiac output which folows from hemodBution. Pv02 measurement, therefore, is a better indicator -19of the oxygenation status of the patient.

Diffusion Oxygen Delivery Oxygen transport to tissue also occurs via diffusion. There are a series of diffusion boundaries through which 02 must pass on its way from the red cell to the tissues. Fick's law of diffusion states that the overal rate of diffusion of a gas from one compartment to another is governed by the diffusion gradient, the difference between the gas concentrations (P 1

-P

2 within the two compartments, and a diffusion constant,

K

d which is a lumped-sum reflection of many factors including properties of the boundary layers, temperature, etc.

d(O 2

K

dt

P-P)

The process of 02 diffusion can be simply illustrated by considering the movement of water through holes in a wall separating a higher elevation reservoir and a lower level reservoir. Water is supplied initialy at one elevation (P 1 and flows to a second lower level (P 2 The hydrostatic pressure driving this movement is the vertical difference in height between the two reservoirs. The total rate of water movement is also limited by the cross-sectional area of the holes in the barrier which provide resistance to flow from compartment 1 to 2 In this analogy, the two water levels correspond to the two 02 pressures (P 1 and P 2 15 in Fick's law of diffusion, shown above, and the cross-sectional area of the holes in the barrier (through which the water flows) would be represented by the diffusion constant, K d Experimental work has shown that there are probably two barriers to diffusion of 02 from the red cal to the tissues: the layer of unstired plasma surrounding the red blood cell and the collective membranes separating the plasma space from the cellular cytosol of adjacent tissue. Raising the PO 2 in the plasma wll 20 have the effect of increasing the rate of diffusion into tissues, since the plasma represents an "intermediate level reservoir" in the preceding analogy. In fact, if there is not a Ianiting supply of 0 2 in red cals, then the rate of movement of 02 from plasma to tissues will be proportional to this plasma reservoir. This represents the essence of the proposed use of low-dose 02 carriers to reduce the need to transfuse alogeneic blood.

The proposed mechanism assumes that a small reduction of the reservoir of available 02 (eg., hemodilution) wil not appreciably change the overall rate of diffusion because it is assumed that the barrier to diffusion represented by the membranes between the plasma and tissue cytosol space is rate-imiting.

Experimental evidence exists to support this assumption.

Increasing the diffusive deivery of 02 to tissue is sometimes caled "diffusion facilitation", and could increase 02 deivery to tissues under conditions where 02 deivery might be otherwise supply-lmited. In other words, increasing the dissolved (plasma) 02 concentration is expected to decrease the level at which critical 02 deivery occurs and thereby increase the margin of safety in terms of prevention of tissue hypoxia.

Experimental evidence suggests that this is, in fact, the case In a study by Faithful Cain U. Cit. Care 3:14-18 (1988)), dogs were initially hemodiluted with either 6% dextran (average molecular weight 70,000, in Tyrode's solution), or the perfluorocarbon emulsion, FLUOSOL and then progressively hemorrhaged to determine the critical 02 extraction ratios. FLUOSOL-treated dogs had lower mixed venous PO2 levels and higher 02 extraction fractions at the critical 02 delivery point. This indicated that perfluorochemicals in FLUOSOL may have promoted diffusion of 02 into the tissues. This effect was very evident in these FLUOSOL studies since these dogs likely had a compromised microcirculation due to the severe capillary flow inhomogeneity that occurs in dogs immediately following injection of only 1 to 2 mL of the FLUOSOL emulsion (Faithful et aL Microvasc. Res. 33:183-93 (1987)).

It should be noted that transfusion of red cells will not affect 02 diffusion in the same manner as described. In fact, an additional physiological effect described by Federspiel et aL Wkrovasc. Res. 32:164-89 (1986)), refers to the fact that in normal capillary beds, red cas are separated by considerable distances as they individually traverse the capilary network. The 02 would be expected to transfer from red cells to tissue predominantly across the area where the red cal is closely in contact with the endothelial cels fining the vasculature. Addition of a cell-free 02 carrier might increase the rate of 02 transfer, simply on the basis that more 02 would be in contact with the endothelial cells.

In general improvement of blood fluidity by hemodiution has been shown to increase mean tissue

PO

2 in various organs (Messmer at aL Res. Exp. Mad. 159:152-56 (1973)). This increase in tissue PO2 was attributed to more even flow distribution at the microcirculatory level and was interpreted as improved tissue oxygenation. On the other hand, Homer Microsc. Res. 22:308-23 (1981), argued that in acute anemia 20 there may be large differences between red blood cell P02 and the plasma P0 2 This would occur as a result of 0 2 diffusion from the red cell being slowed by passage through the plasma (which has very low 02 solubity characteristics). With hemodiution, the spacing between red blood cals in tissue capilaries is increased so that outward diffusion of 02 from red cells is slowed further by the increased diffusional barrier of plasma. The resultant gradient for PO2 may not be resolved not al the oxygen has time to unload) during the short time that the red cell dwels in the capillary and 02 extraction may be diminished accordingly (Gutierrez, Raspikt. Physuil 63:79-96 (1985)) The presence of an additional 02 carrier such as a perfluorochemical in the plasma will increase the total 02 content in the plasma compartment of blood and may faciltate the diffusion of 02 from the red cel into the tissues. The addition of a relatively small dose (3 mL 2.7 g perflubronlkg BW) of a concentrated 90% wlv perflubron emulsion wil result in a significant increase in the total 0 2 content in the plasma. When performed during respiration with 100% 02 and in the presence of acute normovolemic hmnodlution (to a hmatocrit of the net result would represent an increase in the avalable oxygen. Normal oxygen consumption would come preferentially from the perflubron and the plasma, since this 02 is physically dissolved and therefore readily available (compared to the 02 that is chemicaly bound to hemoglobin as a igand). The remaining 02 caried by the red cebs would therefore represnt n avalable reservoi of extra 0 2 that would supply additional oxygen, when needed, to prevent certain sensitive tissues from reaching a P:\dPERAXD\145561l8RSI -17/1100 -21 critical level of 02 delivery.

A low-dose cell-free oxygen carrier is therefore superior, in terms of tissue oxygenation, to additional red cell transfusion. Such an oxygen carrier is used for the temporary enhancement of oxygen delivery during the acute phase of surgery or following organ ischemia or infarct. None of the currently available oxygen carriers can be considered effective "blood substitutes" because of their short retention time in the circulation (hours) compared to red cells (months). With routine use, especially in uncomplicated elective surgery combined with acute normovolemic hemodilution procedures, the "transfusion trigger" can be reduced. With the present invention, wherein PvO 2 or other indices of tissue oxygenation is continuously or periodically monitored and autologous blood or additional oxygen carrier administered to the patient in response to PvO, levels, the "transfusion trigger" can be reduced even further. This can eliminate the need for transfusion of allogenic red blood cells in many cases and thereby significantly reduce the risk of transfusion-bore disease and transfusion reaction. The present invention also provides for hemodilution as an adjunctive therapy for organ ischemia or infarct, by maintaining adequate delivery of oxygen to the tissues while 15 reducing the number of cells known to exacerbate the effects of ischemia and infarct.

EXAMPLE 1 Enhancement of 0, Delivery By Perfluorocarbon Emulsion Immediately prior to undergoing surgery, a patient is subjected to perioperative isovolemic hemodilution. The removed blood is stored for later use. Blood is removed with the concomitant intravenous replacement by a crystalloid solution. During this time, the patient's fractional inspired oxygen concentration (FiO 2 is increased to 1. The patient is hemodiluted until the hemoglobin concentration reaches 8 gm/dL, with each aliquot of the removed blood being replaced by 3 volumes of Ringers-Lactate. A 90% w/v perflubron emulsion having the composition of Formula 1 is administered intravenously to a total dose of 1.8 gm/kg body weight, 2. 5 while the patient's PvO 2 is monitored using a Swan-Ganz catheter. Hemodilution and administration of perflubron emulsion is continued until the PvO, reaches 40 mm Hg (hemoglobin level is 2 gm/dL). Surgery is then initiated, with an attendant blood loss of up to 3 liters.

Autologous blood is then re-administered to the patient to maintain the PvO 2 at 40 mm Hg or above.

Throughout this specification and the claims which follow, unless the context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps.

Although the invention has been described with reference to particular preferred embodiments, the scope of the invention is defined by the following claims and should be construed to include reasonable equivalents.

Claims (7)

1. Use of a biocompatible liquid, a biocompatible oxygen carrier and a patient's removed and stored blood for the manufacture of a medicament for facilitating autologous blood use by said patient when facing a loss of blood, wherein said biocompatible liquid is in a form suitable for intravenous administration in a quantity appropriate to bring the patient's blood hemoglobin level to a desired concentration, said biocompatible oxygen carrier is in a form suitable for intravenous administration in a quantity appropriate to bring the patient's tissue to a desired tissue oxygenation level, and wherein the stored blood is in a form suitable for intravenous administration in a quantity appropriate to maintain oxygenation assessments at or above a desired value.

2. Use of a biocompatible liquid and a biocompatible non-red cell oxygen carrier for the manufacture of a medicament for the treatment of organ ischemia or infarct in a patient, wherein said biocompatible liquid is in a form suitable for intravenous administration in a quantity appropriate to reduce ths patient's blood hemoglobin level to a desired concentration, and said oxygen carrier is in a form suitable for intravenous administration in a quantity appropriate to maintain oxygenation of the 15 patient's tissue at or above a predetermined level in conjunction with removal of blood from said patient.

3. Use of a biocompatible oxygen carrier for the manufacture of a medicament for hemodiluting a patient's blood wherein said biocompatible oxygen carrier is in a form suitable for intravenous administration in a quantity appropriate to maintain oxygenation of the patient's tissue at or above a predetermined level in conjunction with removal of blood from said patient, while .o 20 simultaneously assessing the patient's tissue oxygenation level to provide an oxygenation index reflective of actual tissue oxygenation and additional oxygen carrier is administered to said patient to maintain said oxygenation index at or above a desired value.

4. Use of a biocompatible liquid for the manufacture of a medicament for facilitating autologous blood use by a patient facing a loss of blood in which a portion of the patient's blood is removed and stored while the biocompatible liquid is administered intravenously in sufficient quantity to bring the patient's blood hemoglobin level to a desired concentration; and in which a biocompatible oxygen carrier is administered intravenously, while simultaneously assessing the patient's tissue oxygenation through measurement of a physiological parameter to provide an oxygenation index reflective of actual tissue oxygenation, after which said patient undergoes a further loss of blood; and in which the stored blood is administered intravenously to said patient to maintain said oxygenation index at or above a desired value. .23- Use of a biocompatible liquid for the manufacture of a medicament for the treatment of organ ischemia or infarct, in which a portion of the blood of a patient in need of treatment for organ ischemia or infarct is removed and the biocompatible liquid is administered intravenously in sufficient quantity to reduce the patient's blood hemoglobin level to a desired concentration; and in which a biocompatible non-red cell oxygen carrier is administered in conjunction with said removal of blood to maintain oxygenation of the patient's tissue at or above a predetermined level,

6. Use of a biocompatible liquid, a biocompatible oxygen carrier and a patient's removed and stored blood for facilitating autologous blood use by said patient when facing a loss of blood, wherein said biocompatible liquid is-ina-form suitable for intravenous administration in a quantity appropriate to bring the patient's blood hemoglobin level to a desired concentration, said biocompatible oxygen carrier is in form suitable for intravenous administration in a quantity appropriate to bring the patient's tissue to a desired tissue oxygenation level, and wherein the stored blood is in a form suitable for intravenous administration in a quantity appropriate to maintain oxygenation assessments at or above a desired value. 15

7. Use of a biocompatible liquid and a biocompatible non-red cell oxygen carrier for the treatment of organ ischemia or infarct in a patient, wherein said biocompatible liquid is in a form suitable for intravenous administration in a quantity appropriate to reduce the patient's blood hemoglobin level to a desired concentration, and said oxygen carrier are in form suitable for intravenous administration in a quantity appropriate to maintain oxygenation of the patient's tissue at or above a

20- predetermined level in conjunction with removal of blood from said patient. 8. Use of a biocompatible oxygen carrier for hemodiluting a patient's blood wherein said biocompatible oxygen carrier is in a form suitable for intravenous administration in a quantity appropriate to maintain oxygenation of the patient's tissue at or above a predetermined level in conjunction with :0 removal of blood from said patient. 25 9. Use of a biocompatible liquid for facilitating autologous blood use by a patient facing a loss of blood in which a portion of the patient's blood is removed and stored while the biocompatible liquid is administered intravenously in sufficient quantity to bring the patient's blood hemoglobin level to a desired concentration; and in which a biocompatible oxygen carrier is administered intravenously, while assessing the patient's tissue oxygenation, after which said patient undergoes a further loss of blood; and in which the stored blood is administered intravenously to said patient in response to said oxygenation assessments to maintain oxygenation assessments at or above a desired value. -24- Use of a biocompatible liquid for treatment of organ ischemia or infarct, in which a portion of the blood of a patient in need of treatment for organ ischemia or infarct is removed and the biocompatible liquid is administered intravenously in sufficient quantity to reduce the patient's blood hemoglobin level to a desired concentration; and in which a biocompatible non-red cell oxygen carrier is administered in conjunction with said removal of blood to maintain oxygenation of the patient's tissue at or above a predetermined level. 11. Use of a biocompatible oxygen carrier for hemodiluting a patient, wherein a portion of the patient's blood is removed and stored while the biocompatible oxygen carrier is administered intravenously and the patient's tissue oxygenation is asseused and after which the patient undergoes a further loss of blood. 12. Use according to Claim 1, 2, 4, 5, 6, 7, 9 or 10 wherein the biocompatible liquid further comprises a hemodiluent. 13. Use according to Claim 12, wherein said hemodiluent is administered separately from said oxygen carrier. 15 14. Use according to Claim 1, 4, 6 or 9, wherein additional oxygen carrier is administered in response to said oxygenation assessments to maintain said oxygenation assessments at or above a desired value prior to readministering said stored blood. Use according to any one of Claim 1-11, wherein the oxygen carrier is derived from human, animal, plant, or recombinant hemoglobin. 9n 16. Use according to any one of Claims 1-11, wherein the oxygen carrier is a fluorocarbon r6.**e a 0* a 0 emulsion. 0O 5 a.. 0@SS S S. es a 0@ u £5 1: I: ,hrn said flunrncarbon emulsion has a concentration of 17. Use accoroing to Claii w.1 at least 40%, wiv. 18. Use according to Claim 16, wherein the concentration of said fluorocarbon emulsion is at least 60%, wlv. 19. Use according to Claim 1, 2, 4, 5, 6, 7, 9 or 10, wherein the oxygen carrier is a fluorocarbon emulsion and the volume of said administered oxygen carrier is less than 50% of the volume of said biocompatible liquid. Use according to Claim 1, 2, 4, 5, 6, 7, 9 or 10, wherein said biocompatible liquid is at least one of a crystalloid, colloid, and a biocompatible oxygen carrier. DATED this 23rd day of June, 2000 Alliance Pharmaceutical Corp. By DAVIES COLLISON CAVE Patent Attorneys for the Applicants