EP0290252A2
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EP
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Pasteurizable, freeze-driable hemoglobin-based blood substitute
Assignee: Agriculture and Agri-Food Canada
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Filing: 1988-05-05
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Granted: —
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Expires: 2008-05-05
Abstract
Abstract The present specification describes a process by which a blood substitute (hereinafter referred to as "HemoSafe") is derived from uniformly stabilized monomers and polymers of deoxyhemoglobin in its tight (T) conformation, with oxygen affinity similar to that of human blood. Two classes of HemoSafe are derived respectively from animal-hemoglobin and human-hemoglobin. HemoSafe (animal) differs from HemoSafe (human) in that it is free of polymers in order to reduce potential immunogenicity if used in man. Both types of HemoSafe may be derived in the following manner. The stabilized deoxyhemoglobins are converted to their carbonmonoxy derivatives (CO-HemoSafe) which are then stable under pasteurization conditions to render them viral disease transmission-free. CO-HemoSafes are stable for 2 months at 56°C in either the solution or the freeze-dried state. For transfusion CO-HemoSafes are easily oxygenated under sterile conditions by photoconversion yielding oxy-HemoSafe. In addition a transfusable met-hemoglobin derivative for treatment of cyanide poisoning, is derived by converting oxy-HemoSafe to met-HemoSafe.
Inventors
- 👤 Majesty The Queen In Right Of Canada As Represented By The Minister Of National Defence Of Her Majesty's Canadian Gov. Her
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-11-10 01:57
CPC Classifications
C; C07; C07K; C07K14/00; C07K14/795; C07K14/805; A; A61; A61K; A61K38/00; A61K38/16; A61K38/41; A61K38/42; A61L; A61L2/00; A61L2/02; A61L2/04; A61P; A61P7/00; A61P7/08; A61L2103/00; A61L2103/05
Claims Analysis
Total Claims: 31
Claim 1
Independent Claim
48 words
A hemoglobin product consisting essentially of tetrameric hemoglobin units stabilized against dissociation into dimers, and stabilized against conformational change between the T-conformation and the R-conformation upon formation of aqueous preparations thereof, said tetrameric units having covalent chemical linkages between globin chains of the sub-units to effect the stabilization.
Claim 2
Dependent Claim
48 words
A hemoglobin product consisting essentially of tetrameric hemoglobin units stabilized against dissociation into dimers, and stabilized against conformational change between the T-conformation and the R-conformation upon formation of aqueous preparations thereof, said tetrameric units having covalent chemical linkages between globin chains of the sub-units to effect the stabilization.
Claim 3
Dependent Claim
14 words
Stabilized hemoglobin product according to Claim 1 which is conformationally stabilized into the T-conformation.
Claim 4
Dependent Claim
22 words
Stabilized hemoglobin product according to Claim 2, wherein the hemoglobin is stabilized by secondary amino linkages between globin chains of the hemoglobin.
Claim 5
Dependent Claim
33 words
Stabilized hemoglobin product according to Claim 3, wherein the links are derived from reaction of di- or polyaldehydes with globin chains to form Schiff base links followed by reduction to secondary amino linkages.
Claim 6
Dependent Claim
14 words
Stabilized hemoglobin product according to Claim 4 in which the aldehyde reactant is glutaraldehyde.
Claim 7
Dependent Claim
14 words
Stabilized hemoglobin product according to Claim 4 in which the aldehyde reactant is glyoxal.
Claim 8
Dependent Claim
21 words
Stabilized hemoglobin product according to Claim 4 wherein the aldehyde is the product of ring opening oxidation of a sugar compound.
Claim 9
Dependent Claim
20 words
Stabilized hemoglobin product according to Claim 7 wherein the sugar compound has from 1 - 7 sugar residues per molecule.
Claim 10
Dependent Claim
13 words
Stabilized hemoglobin product according to Claim 8 wherein the sugar compound is raffinose.
Claim 11
Dependent Claim
14 words
Stabilized hemoglobin product according to Claim 1 wherein the hemoglobin is of human origin.
Claim 12
Dependent Claim
14 words
Stabilized hemoglobin product according to Claim 1 wherein the hemoglobin is of animal origin.
Claim 13
Dependent Claim
95 words
A process for preparing a stabilized hemoglobin product according to Claim 1, which comprises reacting hemoglobin in aqueous solution with a divalent or polyvalent reagent bearing at least a pair of functional groups capable of room temperature reaction with at least two respective groups or globin chains of the hemoglobin, said reagent being free from negatively charged ionic groups capable of reacting with the DPG site of the hemoglobin; deactivating residual functional groups on said reagent after the reaction with hemoglobin is substantially complete; and recovering the conformationally stabilized and dissociatively stabilized hemoglobin so formed.
Claim 14
Dependent Claim
21 words
The process of Claim 12 wherein said reagent has functional groups capable of reaction with amino groups or the globin chains.
Claim 15
Dependent Claim
22 words
The process of Claim 13 wherein said reagent is a dialdehyde or a polyaldehyde, and its residual groups are deactivated by reduction.
Claim 16
Dependent Claim
18 words
The process of Claim 13 wherein the hemoglobin reactant and the stabilized hemoglobin product are in the T-conformation.
Claim 17
Dependent Claim
37 words
A stabilized hemoglobin product comprising a plurality of said stabilized tetrameric hemoglobin units according to Claim 1, covalently linked together to form a polymeric hemoglobin product in which each hemoglobin unit has conformational stability and dissociative stability.
Claim 18
Dependent Claim
19 words
A stabilized polymer hemoglobin product according to Claim 16, wherein each hemoglobin unit thereof is stabilized in the T-conformation.
Claim 19
Dependent Claim
33 words
A stabilized polymeric hemoglobin product according to Claim 17, wherein the covalent links between hemoglobin units comprise secondary amine linkages derived from dialdehyde or polyaldehyde reaction with the stabilized hemoglobin followed by reduction.
Claim 20
Dependent Claim
65 words
A stabilized polymeric hemoglobin product according to Claim 17 and having a composition such that from 20 - 25% of the constituents are hemoglobin polymers consisting of five or more hemoglobin units, 50 - 60% of the constituents are hemoglobin oligomers of 2 - 4 hemoglobin units, 20 - 25% of the constituents are monomeric hemoglobin, and 1 - 2% are dimers of hemoglobin subunits.
Claim 21
Dependent Claim
34 words
A stabilized hemoglobin product according to Claim 1, complexed with carbon monoxide, said complex in aqueous solution being capable of withstanding temperatures of up to 80°C for 10 hours without substantial denaturation or decomposition.
Claim 22
Dependent Claim
17 words
A stabilized CO-hemoglobin complex according to Claim 20 wherein the hemoglobin units are stabilized in their T-conformation.
Claim 23
Dependent Claim
11 words
A stabilized CO-hemoglobin complex according to Claim 21 in lyophilized form.
Claim 24
Dependent Claim
16 words
A stabilized CO-hemoglobin complex according to Claim 21 wherein the CO:hemoglobin molar ratio is about 4:1.
Claim 25
Dependent Claim
74 words
A process of preparing a hemoglobin product suitable for use as a blood substitute and free of transmissible viral infection, which comprises pasteurizing the carbon monoxide-stabilized hemoglobin complex of Claim 21 by warming a solution thereof to appropriate pasteurization temperatures for times sufficient to cause effective deactivation of any viral contaminants of said complex and subsequently reacting the complex with oxygen to convert it to an oxy- or deoxy-hemoglobin complex, or a mixture thereof.
Claim 26
Dependent Claim
21 words
Pasteurized, viral inactivated blood substitute containing conformationally and dissociatively stabilized oxy- or deoxy-hemoglobin or a mixture thereof, according to Claim 1.
Claim 27
Dependent Claim
28 words
A process of preparing met-hemoglobin complex suitable for use as a blood substitute for cyanide scavenging which comprises oxidizing the pasteurized oxyhemoglobin of Claim 25, under appropriate conditions.
Claim 28
Dependent Claim
56 words
A biological proteinaceous substance comprising macromolecules capable of existing in aqueous solution in a plurality of different conformations, at least 95% of said macromolecules being stabilized into one specific conformation, said stabilization having been effected by non-site specific reaction of said macromolecules with a dialdehyde or polyaldehyde, followed by reduction to form covalent secondary amino linkages.
Claim 29
Dependent Claim
56 words
A biological proteinaceous substance comprising macromolecules capable of existing in aqueous solution in a plurality of different conformations, at least 95% of said macromolecules being stabilized into one specific conformation, said stabilization having been effected by non-site specific reaction of said macromolecules with a dialdehyde or polyaldehyde, followed by reduction to form covalent secondary amino linkages.
Claim 30
Dependent Claim
76 words
A process for pasteurization of a hemoglobin based blood substitute product to render the product essentially free of viral contamination, which comprises reacting said blood substitute with carbon monoxide to protect the heme against ferrous-ferric oxidation, maintaining the carbon monoxylated blood substitute at appropriate pasteurization temperatures for times sufficient to cause effective deactivation of viral contaminants, and subsequently reacting the complex with oxygen to convert it to an oxy- or deoxy-hemoglobin product, or a mixture thereof.
Claim 31
Dependent Claim
76 words
A process for pasteurization of a hemoglobin based blood substitute product to render the product essentially free of viral contamination, which comprises reacting said blood substitute with carbon monoxide to protect the heme against ferrous-ferric oxidation, maintaining the carbon monoxylated blood substitute at appropriate pasteurization temperatures for times sufficient to cause effective deactivation of viral contaminants, and subsequently reacting the complex with oxygen to convert it to an oxy- or deoxy-hemoglobin product, or a mixture thereof.
References Cited (Backward Citations)
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