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WO2018172422A1 - Method of preserving erythrocytes using pvac - Google Patents

Method of preserving erythrocytes using pvac Download PDF

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Publication number
WO2018172422A1
WO2018172422A1 PCT/EP2018/057200 EP2018057200W WO2018172422A1 WO 2018172422 A1 WO2018172422 A1 WO 2018172422A1 EP 2018057200 W EP2018057200 W EP 2018057200W WO 2018172422 A1 WO2018172422 A1 WO 2018172422A1
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WO
WIPO (PCT)
Prior art keywords
group
composition
groups
carrier
less
Prior art date
Application number
PCT/EP2018/057200
Other languages
French (fr)
Inventor
Thomas Engstrand
David Berglund
Original Assignee
Pvac Medical Technologies Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Pvac Medical Technologies Ltd filed Critical Pvac Medical Technologies Ltd
Publication of WO2018172422A1 publication Critical patent/WO2018172422A1/en

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/74—Synthetic polymeric materials
    • A61K31/785—Polymers containing nitrogen
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N1/00—Preservation of bodies of humans or animals, or parts thereof
    • A01N1/10—Preservation of living parts
    • A01N1/12—Chemical aspects of preservation
    • A01N1/122—Preservation or perfusion media
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N1/00—Preservation of bodies of humans or animals, or parts thereof
    • A01N1/10—Preservation of living parts
    • A01N1/14—Mechanical aspects of preservation; Apparatus or containers therefor
    • A01N1/146—Non-refrigerated containers specially adapted for transporting or storing living parts whilst preserving
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61J—CONTAINERS SPECIALLY ADAPTED FOR MEDICAL OR PHARMACEUTICAL PURPOSES; DEVICES OR METHODS SPECIALLY ADAPTED FOR BRINGING PHARMACEUTICAL PRODUCTS INTO PARTICULAR PHYSICAL OR ADMINISTERING FORMS; DEVICES FOR ADMINISTERING FOOD OR MEDICINES ORALLY; BABY COMFORTERS; DEVICES FOR RECEIVING SPITTLE
    • A61J1/00—Containers specially adapted for medical or pharmaceutical purposes
    • A61J1/05—Containers specially adapted for medical or pharmaceutical purposes for collecting, storing or administering blood, plasma or medical fluids ; Infusion or perfusion containers
    • A61J1/10—Bag-type containers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
    • A61K35/14—Blood; Artificial blood
    • A61K35/18—Erythrocytes

Definitions

  • the present invention relates to the use of a composition for stabilizing and preserving erythrocytes or red blood cells.
  • Plastic bag systems for whole blood collection, component separation, and storage are required for current blood banking practice and transfusion therapies. Bags consisting of different plastic materials are commercially available but polyvinyl chloride (PVC) plasticized with di-(2-ethylhexyl) phthalate (DEHP) is the most widely used material.
  • PVC polyvinyl chloride
  • DEHP di-(2-ethylhexyl) phthalate
  • DEHP-containing devices relate to: (1) the aggregate exposure and (2) the sensitivity of the exposed patient population. While devices used in neonates deserve particular attention, there may be other patient subgroups where DEHP exposure may be anissue. Regulatory agencies therefore urge minimizing patient exposure to DEHP and recommend considering replacing DEHP with alternative materials. However, in several circumstances, e.g. in blood preservation, such replacements are currently not available. Blood bags containing DEHP tend to release DEHP into the blood. Even though DEHP stabilize the membrane of the erythrocytes and reduce haemolysis there is a need for an alternative to DEHP that is not toxic.
  • the present inventors have recognized that immobilization of relevant active functionalities to a macro molecular carrier improves the effects of the corresponding low molecular weight active molecule, forming an active principle. Unexpectedly it has been found that the optimal functionalities have been found amongst reactive groups that normally are used for cross-linking macromolecular carriers and/or for the synthesis of carrier-bound therapeutic active entities (WO 2009108100 and references cited therein).
  • FIG. 1 graph disclosing the effect of the present invention on hemolysis for different concentrations of the active principle (PVAC).
  • K denotes control (PBS).
  • the first aspect is a composition for use in preserving and stabilizing erythrocytes.
  • AP active principle
  • the second main aspect of the invention is a method for preserving or stabilizing
  • the main characteristic feature of the method comprises the steps of:
  • a mixture comprising erythrocytes and the composition according to the present invention.
  • a container having an inner and an outer surface wherein the inner surface is coated with the composition or active principle according to the present invention FIFTH MAIN ASPECT (coated tube)
  • a tube such as a dialysis tube having an inner and an outer surface wherein the inner surface is coated with the composition or active principle.
  • the composition may contain one or more formulations where at least one of them comprises the AP or reagents necessary for the formation in vivo of AP (e.g. as described in WO 2009108100 for compositions used for formation in vivo of extracellular matrices).
  • the AP comprises a carrier exhibiting the plurality of active structures. Every active structure is firmly attached to the carrier, for instance covalently.
  • AP as well as the carrier as such may be soluble or insoluble in aqueous liquids such as water, body fluids, such as blood, serum, plasma, urine, lymph, lachrymal fluid, intestinal juice, gastric juice, saliva, synovial fluid etc.
  • the AP may be fixed to a water-insoluble support that may be of various physical and/or geometric appearances depending on the intended use,see further below.
  • Either one or both of the carrier and the support should be inert in the sense that they should not participate in the preserving or stabilizing activity or reaction. Both of them should have an acceptable biocompatibility causing low or no host defence reactions including low or no inflammation.
  • the active structure comprises a first nucleophilic centre.
  • the preferred heteroatoms are N and S.
  • S is preferably combined with the presence of a second nucleophilic centre, such as a primary or secondary amino, in the same active 5 structure as discussed below.
  • the bivalent organic group -R"- provides binding to the carrier via one of its free valences.
  • the other free valency of -R' '- as well as the free valency of the other organic group R'- are directly attached to the heteroatom X 1 .
  • nucleophilic centre has the formula:
  • a single-bonded atom means that the atom is directly bound to other atoms only by single 15 bonds.
  • a multiple-bonded atom means that the atom is directly bound to another atom by a triple or a double bond.
  • the atoms referred to are primarily N, O, S and carbon.
  • nucleophilic centres are typically uncharged.
  • Either one or both of the organic groups R'- and -R"- comprise a structure of the formula
  • each of m', n' and o' is 0 or 1, with preference for m' being 1 with further preference for either one or both of n' and o' also being 1,
  • each of X 2 , X 3 , and X 4 is selected amongst NH and a heteroatom S or O, with
  • the left free valence provides binding to a monovalent alkyl group R*- or to the carrier via at least a bivalent alkylene group -R**-, each of which comprises the methylene group -CH 2 - shown of formula II,
  • Either one or both of the monovalent alkyl group R*- and the bivalent alkylene -R**- may be straight, branched or cyclic and possibly contain one or more structures selected amongst ethers (-0-, -S-), hydroxy (-OH), mercapto (-SH) and amino (-NH-, -NH 2 ).
  • Either one or both of these alkyl groups are preferably a lower alkyl which in this context means that they comprise one, two, three, four, five up to ten sp 3 -hybridised carbons typically with at most one heteroatom O, N and S bound to one and the same carbon.
  • the hydrogens given in formula (I) and/or its substructures may be replaced with an alkyl group selected amongst the same alkyl groups as discussed for R*-.
  • the bivalent group -R"- which attaches the first nucleophilic centre to the carrier comprises a substructure complying with formula I and/or II.
  • Preferred active structures thus have a nucleophilic centre which contain the first heteroatom X 1 together with a structure complying with formula II and comprises a group selected amongst:
  • hydrazide groups such as -NH-NH 2 , e.g. as part of a -CONHNH 2 group, a semicarbazide group such as -NHCONHNH 2 , a carbazate group such as -OCONHNH 2 , a thiosemicarbazide group such as -NHCSNHNH 2 , a thiocarbazate group such as -OCSNHNH 2 ,
  • the free valence indicated in each of the groups given in the preceding paragraph preferably attaches the nucleophilic centre to the carrier via a linker structure comprising the above- mentioned bivalent alkylene group -R**-.
  • a hydrogen bound directly to nitrogen may be replaced with a monovalent alkyl group selected amongst the same alkyl groups as R*- as long as they are not substantially counteracting the desired reactivity of the unsubstituted form of the nucleophilic centre.
  • R*- monovalent alkyl group selected amongst the same alkyl groups as R*- as long as they are not substantially counteracting the desired reactivity of the unsubstituted form of the nucleophilic centre.
  • the hydrogen in a thiol group and in a hydroxyl group cannot be replaced, for instance.
  • Two replacing alkyl groups may form a cyclic structure together with atom to which they are attached, i.e. form a bivalent alkylene group e.g.
  • bivalent structures -R**- and -R"- discussed above comprises next to the carrier a linker structure which does not negatively affect the desired effect of the nucleophilic centre of the active structure.
  • Such structures are not part of the invention and suitable such structures can be designed by the average-skilled person in the field.
  • this second nucleophilic centre complies with the formula:
  • the second nucleophilic centre comprises a group selected amongst:
  • amino groups preferably primary or secondary amino groups
  • hydrazide groups such as -NH-NH 2 , e.g. as part of a -CONHNH 2 group, a semicarbazide group such as -NHCONHNH 2 , a carbazate group such as -OCONHNH 2 , a thiosemicarbazide group such as -NHCSNHNH 2 , a thiocarbazate group such as -OCSNHNH 2 ,
  • thiol groups e.g.-SH.
  • the heteroatom Y 1 preferably is part of
  • the distance between the first heteroatom Y 1 and the first heteroatom X 1 is typically larger than two or three atoms with upper limits being e.g. 20 atoms with preference for 4, 5 or 6 atoms between these two heteroatoms.
  • the distance should support intra-molecular cyclisation, typically via one or more addition reactions.
  • suitable carriers depends on the requirements of a particular use.
  • the typical carrier is selected amongst macro molecular compounds, i.e. is a compound which has a molecular weight of > 2000 dalton, preferably > 10000 dalton or > 50000 dalton, and preferably exhibits a polymeric structure, i.e. is a polymer which may be a homopolymer, copolymer or a chemical adduct between two or more polymers of different polymeric structure.
  • Other suitable carriers may have molecular weights ⁇ 2000 dalton and
  • duct polymer in this context means a product formed by reacting two polymers exhibiting mutually reactive groups capable of forming covalent bonds that link the two polymers together upon reaction of the two mutually reactive groups with each other. See for instance WO 2009108100 (IPR-Systems AB) and references cited therein.
  • a carrier polymer may be cross- linked or not cross-linked. With respect to branching the polymer may be unbranched, i.e. linear, or branched including either hyperbranched or dendritic.
  • the degree of branching may thus vary between 0 and 1, such as be > 0.10 or > 0.25 > 0.5 > 0.75 or > 0.90 and/or ⁇ 0.90 or ⁇ 0.75 or ⁇ 0.50 or ⁇ 0.25 or ⁇ 0.10.
  • Cross-linked polymers are as a rule insoluble in aqueous liquids while the solubility of non-cross-linked polymers depend on the overall structure of the polymer, e.g. presence and amount of polar and/or hydrophilic groups.
  • Carrier polymers may also be derivatized to contain non-polymeric or polymeric groups, for instance cross-links, substituents, charged or uncharged groups, active structures (as discussed above) etc.
  • Macromolecular carriers which are insoluble in aqueous liquids may have different physical and geometric shapes as discussed for support materials elsewhere in this specification.
  • polymer above includes organic as well as inorganic polymers.
  • the macro molecule or polymer used in the carrier may be water-insoluble and suspended in aqueous liquid media (when in particle form).
  • Polymers and other macromolecules suitable as carrier material may be hydrophilic or hydrophobic with preference for hydrophilic.
  • the introduction of hydrophilic groups may among others be accomplished by:
  • Hydrophilic groups/compounds typically have an r > 0.5, preferably > 1.0, and for hydrophobic groups r ⁇ 1.0, preferably ⁇ 0.5.
  • Typical hydrophilic groups are hydroxy, amino, amido, carboxy (including free acid carboxyl as well as carboxylate (ester ester and salt) etc.
  • Typical hydrophobic groups are alkyls (C n H( 2n +i)-, C n H( 2n _i)-, C n H( 2n _3)- etc), phenyls including alkyl phenyls, benzyl including other phenylalkyls etc.
  • a carrier macromolecule typically comprises a polymer backbone which comprises > 5, or more preferably > 10 such as > 25 different and/or identical monomeric units linked together. The polymer may carry projecting or pending polymeric and/or non-polymeric groups of various lengths and kinds.
  • a carrier polymer is preferably hydrophilic with hydrophilic groups selected amongst those given elsewhere in this specification.
  • the most preferred hydrophilic group is hydroxy with the preferred carrier polymers and/or other macro molecular carrier being selected by poly hydroxy polymers (PHP or PH-polymers) exhibiting > 5, with preference for > 10, such as > 25 or > 50 hydroxyl groups and/or > 5 monomeric subunits each of which exhibits one, two, three, four or more hydroxyl groups per unit.
  • PGP poly hydroxy polymers
  • Typical polymers that may be present in polymeric carriers are a) polyester polymers, b) polyamide polymers, c) polyether polymers, d) polyvinyl polymers, e) polysaccharides etc.
  • a carrier may comprise one or more of these polymers/polymeric structures.
  • Polyester polymers are in particular obtained by polymerisation of a) monomers exhibiting at least one hydroxy group and at least one carboxy group, or b) a mixture containing monomers exhibiting two or more hydroxy groups and monomers exhibiting two or more carboxy group.
  • Polyamide polymers are in particular obtained by polymerisation of a) monomers exhibiting at least one amino group and at least one carboxy group, or b) a mixture containing monomers exhibiting two or more amino groups and monomers exhibiting two or more carboxy group.
  • polyamides are those that exhibit polypeptide structure together with a plurality of hydroxy groups (PH-polymers).
  • Suitable polyamide polymers of this kind are typically based on hydroxy-,amino-carboxylic acids as monomers, in particular with the amino group positioned a to the carboxylic group, e.g. serine, threonine, tyrosine, proline etc.
  • Polyether polymers are typically used in combination with other polymeric structures, e.g. polymers of (a), (b), (d) and/or (e) above, which are polyfunctional with respect to the presence of groups such as hydroxy, amino etc.
  • Typical polyether polymers are polyethylene oxide and various copolymerisates between ethylene oxide and other lower alkylene oxides, lower epihalohydrins etc.
  • Polyvinyl polymers which may be suitable as polymeric carriers in the invention are typically found amongst polymers containing one, two or more different monomeric units selected amongst hydroxyalkyl acrylates and methacrylates, N-hydroxyalkyl acryl- and N- hydroxyalkyl methacrylamides, hydroxyalkyl vinyl ethers, vinyl esters etc. In one
  • the carrier is a polyvinyl alcohol (PVA).
  • PVA polyvinyl alcohol
  • Polyvinyl alcohols are typically obtained by partial hydrolysis of polyvinyl esters meaning that polyvinyl alcohols that are preferred in the invention typically exhibit residual amounts of ester groups ( ⁇ 10 % or ⁇ 5%).
  • Typical polysaccharides that may be present in carriers used in the invention include dextran, starch, agarose, agaropektin, cellulose, glucosamino glucanes (GAG), and derivates of these polysaccharides etc.
  • the most interesting polysaccharides are dextran, certain glucosamino glucanes (GAG) such as hyaluronic acid etc.
  • a polymer to be used in the carrier may have been derivatized, e.g. cross-linked and/or functionalized after its synthesis.
  • the active structure including the first, the optional second nucleophilic centre and the various heteroatoms discussed for the active structures are typically part of one and the same 20 organic group/substituent attached to the macromolecular carrier. In certain variants different parts of an active structure may be part of different groups/substituents attached to the carrier and/or part of the carrier.
  • suitable carrier polymers will among others depend on the actual 25 application/use of the composition/method of the invention.
  • suitable polymeric carriers with respect to a particular polymeric structure and/or size may vary within a wide interval.
  • the number of monomeric subunits (mean value) of a polymer present in the carrier may be > 20 or > 100 or > 200 or > 300 or > 500 or > 1000 or > 2000 or > 20 000 or > 50 000 and/or ⁇ 50 000 or ⁇ 20 000 or ⁇ 2000 or ⁇ 1000 or ⁇ 500 or ⁇ 300 or ⁇ 200, or ⁇ 30 100 (with the proviso that >-limit always is lower than a ⁇ - limit when these values are
  • Suitable numbers of monomeric units may in some cases be found in the interval of 200 - 600 which in particular applies to the polyvinylalcohol used in the experimental part.
  • Suitable numbers of active structures or nucleophilic centres per monomeric unit of a polymer of the carrier will also depend on the use, the active structure, etc and may thus be found within a wide interval, such as ⁇ 80 %, such as ⁇ 50% or ⁇ 20 % with typical lower limits being 0.01 % or 0.1 % or 1 % where 100% corresponds to one active structure or nucleophilic centre per monomeric unit.
  • For active structures containing two or more nucleophilic centres the number of nucleophilic centres per monomeric unit may exceed 100 %, such as > 100% or > 125 % or > 150%.
  • the AP is present in the composition as an AP-formulation in which the AP is:
  • dissolved in this context means that the AP is present as a solute.
  • the AP particles comprise the AP in a pure form or diluted with some solid material. Useful concentrations of the AP in formulations according to (b) can be found within a broad interval.
  • composition may in addition to the AP contain an aqueous solution such as a buffer (e.g. PBS buffer), saline solution, salts etc.
  • a buffer e.g. PBS buffer
  • saline solution e.g. glycerol, glycerol, glycerol, glycerol, glycerol, glycerol, glycerol, glycerol, saline solution, salts etc.
  • active principle may more or less be the only component of the composition.
  • the AP may be synthesized according to well-known protocols, for instance of the kinds given in WO 2009108100 (IPR-Systems AB) and references cited therein.
  • Water-insoluble support for instance of the kinds given in WO 2009108100 (IPR-Systems AB) and references cited therein.
  • the AP may be fixed to a water-insoluble support.
  • This support material may be selected amongst support materials that have at least one or more of the following characteristics: a) in the form of particles, b) porous or non-porous particles or monoliths allowing or not allowing, respectively, aqueous liquids to penetrate the support, c) rigid, d) soft, e) elastic, f) compressible, g) gellable (in particular to form a hydrogel when placed in contact with water) etc.
  • the support may comprise plastics, glass, mineral, metal etc.
  • the carrier of the AP When the carrier of the AP is insoluble in aqueous media the carrier as such may define its own 5 solid support.
  • Suitable supports typically comprise polymeric materials, e.g. comprising one or more polymer selected from the same polymers as the carrier polymers are selected.
  • Typical carrier polymers are polysaccharides, e.g. cellulose, cross-linked dextran, agarose such as 10 cross-linked agarose etc, polyester polymers e.g. lactic acid copolymers such as polyglactin, polyethylenes etc.
  • the polymer is polyvinyl chloride (PVC).
  • Other kinds of support material may also be used, e.g. ceramic materials, plastics, mineral materials, metals, composite material, activated carbon etc. Porous forms of theses materials may be used as filters and/or adsorbent material.
  • Attachment to the support may be accomplished by mixing, coating, impregnating etc the support with AP according to techniques known in the field. Alternatively, the
  • macro molecular carrier of AP may be part of the material from which a support/device is made.
  • the contacting of the AP with the erythrocytes may take place in vivo of or separate from the individual to be treated.
  • the amount of the AP in the composition which is brought into contact with the red blood cells (erythrocytes) is effective in the sense that the cells are stabilized or preserved with reduced haemolysis as a result.
  • the suitable dosage (per administration) for in-vivo applications depends partly on formulation (e.g. the kind of support material, AP, , concentrations etc) etc and thus is selected within a broad interval, e.g. 5 ⁇ g/ml or more, or
  • the concentration is 10 ⁇ g/ml to lOmg/ml such as 100 ⁇ g/ml to 1 mg/ml.
  • Containers suitable for containing erythrocytes may contain the AP.
  • the container may be a bag, such as a blood bag, a syringe, a vial, a beaker, an e where the container is coated with the AP according to the invention.
  • the container may be made of any suitable material such as a plastic material for example polyvinyl chloride (PVC).
  • the composition may be applied to the surface of the container by dip, spin or spray coating the surface of the container or using any other suitable technique.
  • the composition may comprise a suitable solvent. Adding a solvent to the composition may make it easier to apply the composition to the surface of the container.
  • the solvent may be an aqueous solvent such as water or a buffer (e.g. PBS buffer), or an alcohol such as ethanol.
  • a tube such as a dialysis tube may also be coated with the composition or AP.
  • the inner surface of the tube facing the erythrocytes may be coated with the AP by applying the composition using any suitable technique as described above.
  • the surface of the support or the device may be pre-treated in order to enhance the attachment or adsorption of the composition or AP.
  • the amount of AP on the surface should be such that the stabilizing or preserving effect of the AP is obtained.
  • the present invention also relates to a mixture of erythrocytes and the composition according to the present invention.
  • the erythrocytes are stabilized or preserved in the mixture and may therefore be stored for a longer time without extensive haemolysis.
  • the mix may comprise 5 ⁇ g/ml or more, or 10 ⁇ g/ml or more, or 20 ⁇ g/ml or more, or 100 ⁇ g/ml or more, or 200 ⁇ g/ml or more, or 500 ⁇ g/ml or more, or 20 mg/ml or less, but preferably 10 mg/ml or less, or 5 mg/ml or less, or 1 mg/ml or less.

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Abstract

Use of a composition for stabilizing erythrocytes wherein the composition comprises as an active principle (= AP) a carrier which exhibits a plurality of active structureactive structures, said active structureactive structures comprising a nucleophilic centre complying with the formula: X1(-R''-)(-R')mHn where a) X1 is a single-bonded heteroatom selected amongst N, O and S and exhibits a free electron pair, b) m is 0 or 1 and n is 1 or 2, c) -R"- is a bivalent organic group providing attachment to the carrier via one of its free valences and to X1 at the other free valence, and d) R'- is a monovalent organic group attached to the X1 via its free valence.

Description

METHOD OF PRESERVING ERYTHROCYTES USING PVAC
TECHNICAL FIELD
The present invention relates to the use of a composition for stabilizing and preserving erythrocytes or red blood cells.
BACKGROUND TECHNOLOGY
Plastic bag systems for whole blood collection, component separation, and storage are required for current blood banking practice and transfusion therapies. Bags consisting of different plastic materials are commercially available but polyvinyl chloride (PVC) plasticized with di-(2-ethylhexyl) phthalate (DEHP) is the most widely used material.
However, the toxicity of DEHP and congener phthalates has been and remains a topic of public concern. For red blood cell (RBC) components, replacing DEHP by a non-phthalate PVC plasticizer is challenged by the membrane protective effect that DEHP exerts on RBCs ex vivo. The enhanced RBC in vivo recovery observed with DEHP underlines the plasticizer' s role in the plasma membrane preservation during storage.
The risks from DEHP-containing devices relate to: (1) the aggregate exposure and (2) the sensitivity of the exposed patient population. While devices used in neonates deserve particular attention, there may be other patient subgroups where DEHP exposure may be anissue. Regulatory agencies therefore urge minimizing patient exposure to DEHP and recommend considering replacing DEHP with alternative materials. However, in several circumstances, e.g. in blood preservation, such replacements are currently not available. Blood bags containing DEHP tend to release DEHP into the blood. Even though DEHP stabilize the membrane of the erythrocytes and reduce haemolysis there is a need for an alternative to DEHP that is not toxic.
OBJECTS
The primary goal with the present invention is to overcome the drawbacks of the prior art. INVENTION
The present inventors have recognized that immobilization of relevant active functionalities to a macro molecular carrier improves the effects of the corresponding low molecular weight active molecule, forming an active principle. Unexpectedly it has been found that the optimal functionalities have been found amongst reactive groups that normally are used for cross-linking macromolecular carriers and/or for the synthesis of carrier-bound therapeutic active entities (WO 2009108100 and references cited therein).
In the present specification an active principle of the above-mentioned structure is shown to preserve and stabilize erythrocytes. So far, the mechanism(s) for the preservation and stabilisation is/are unknown. BRIEF DESCRIPTION OF THE FIGURES
Figure 1, graph disclosing the effect of the present invention on hemolysis for different concentrations of the active principle (PVAC). K denotes control (PBS).
FIRST MAIN ASPECT
The first aspect is a composition for use in preserving and stabilizing erythrocytes. The main characteristic feature is that the composition comprises as an active principle (= AP) comprising a carrier which exhibits a plurality of active structures which when present on the carrier is capable of preserving and stabilizing erythrocytes. SECOND MAIN ASPECT (METHOD)
The second main aspect of the invention is a method for preserving or stabilizing
erythrocytes. The main characteristic feature of the method comprises the steps of:
i) providing a composition according to the present invention, and
ii) contacting the composition with erythrocytes.
THIRD MAIN ASPECT (mixture)
A mixture comprising erythrocytes and the composition according to the present invention.
FOURTH MAIN ASPECT (coated container)
A container having an inner and an outer surface wherein the inner surface is coated with the composition or active principle according to the present invention. FIFTH MAIN ASPECT (coated tube)
A tube such as a dialysis tube having an inner and an outer surface wherein the inner surface is coated with the composition or active principle.
THE COMPOSITION
The composition may contain one or more formulations where at least one of them comprises the AP or reagents necessary for the formation in vivo of AP (e.g. as described in WO 2009108100 for compositions used for formation in vivo of extracellular matrices).
The active principle (= AP)
The AP comprises a carrier exhibiting the plurality of active structures. Every active structure is firmly attached to the carrier, for instance covalently. AP as well as the carrier as such may be soluble or insoluble in aqueous liquids such as water, body fluids, such as blood, serum, plasma, urine, lymph, lachrymal fluid, intestinal juice, gastric juice, saliva, synovial fluid etc.
The AP may be fixed to a water-insoluble support that may be of various physical and/or geometric appearances depending on the intended use,see further below.
Either one or both of the carrier and the support should be inert in the sense that they should not participate in the preserving or stabilizing activity or reaction. Both of them should have an acceptable biocompatibility causing low or no host defence reactions including low or no inflammation.
The active structure
The active structure comprises a first nucleophilic centre. The nucleophilic centre (first centre) of the active structure preferably comprises a single- bonded first heteroatom N, O or S (= X1) which exhibits
a) a free electron pair,
b) one or two hydrogens, and c) one or two organic groups R'- (monovalent) and -R"- (divalent) directly bound to the heteroatom.
The preferred heteroatoms are N and S. S is preferably combined with the presence of a second nucleophilic centre, such as a primary or secondary amino, in the same active 5 structure as discussed below. The bivalent organic group -R"- provides binding to the carrier via one of its free valences. The other free valency of -R' '- as well as the free valency of the other organic group R'- are directly attached to the heteroatom X1.
Generically a nucleophilic centre has the formula:
10 X^-R^X-R mHn (formula I)
where X1 , R'- and -R"- are as defined in the preceding paragraph and m is 0 or 1 and n is 1 or 2 with the sum of m plus n being 2 for X1 = N and 1 for X1 = S and O.
A single-bonded atom means that the atom is directly bound to other atoms only by single 15 bonds. A multiple-bonded atom means that the atom is directly bound to another atom by a triple or a double bond. The atoms referred to are primarily N, O, S and carbon.
The preferred nucleophilic centres are typically uncharged. For a nucleophilic centre which is an uncharged base or acid form of an acid-base pair > 5 %, such as > 25% or > 50 or > 20 75%, of the total concentration of the acid-base pair should be in uncharged form.
Either one or both of the organic groups R'- and -R"- comprise a structure of the formula
-CH2(X4V(C=X3)n>(X2)m>- (formula II)
where
25 a) each of m', n' and o' is 0 or 1, with preference for m' being 1 with further preference for either one or both of n' and o' also being 1,
b) each of X2 , X3, and X4, is selected amongst NH and a heteroatom S or O, with
preference for either one or both of X2 and X4 being selected amongst NH and O with further preference for X3 being selected amongst NH, O and S,
30 c) the left free valence provides binding to a monovalent alkyl group R*- or to the carrier via at least a bivalent alkylene group -R**-, each of which comprises the methylene group -CH2- shown of formula II,
d) the right free valency binds directly to the first heteroatom X1. The substructure C=X3 (= B) includes also other ester- and amide-forming substructures which derive from acid functions and form an ester function when X2 and/or X4 are oxygen and/or an amide function when X2 and/or X4 are NH, e.g. sulphonamide (B is S(=0)2) or phosphone amide (B is P=0(NH2) or P=0(OH), n' = 1).
Either one or both of the monovalent alkyl group R*- and the bivalent alkylene -R**- may be straight, branched or cyclic and possibly contain one or more structures selected amongst ethers (-0-, -S-), hydroxy (-OH), mercapto (-SH) and amino (-NH-, -NH2). Each free valences represent binding to sp3-hybridised carbon (= alkyl carbon). Either one or both of these alkyl groups are preferably a lower alkyl which in this context means that they comprise one, two, three, four, five up to ten sp3 -hybridised carbons typically with at most one heteroatom O, N and S bound to one and the same carbon. The hydrogens given in formula (I) and/or its substructures may be replaced with an alkyl group selected amongst the same alkyl groups as discussed for R*-.
It is preferred that the bivalent group -R"- which attaches the first nucleophilic centre to the carrier comprises a substructure complying with formula I and/or II.
The structural elements (substructures) discussed in the preceding paragraphs will support de localisation of electrons.
Preferred active structures thus have a nucleophilic centre which contain the first heteroatom X1 together with a structure complying with formula II and comprises a group selected amongst:
a) amino groups preferably primary or secondary amino groups
b) hydrazide groups such as -NH-NH2, e.g. as part of a -CONHNH2 group, a semicarbazide group such as -NHCONHNH2, a carbazate group such as -OCONHNH2, a thiosemicarbazide group such as -NHCSNHNH2, a thiocarbazate group such as -OCSNHNH2,
c) aminooxy groups, such as -ONH2 etc,
d) a thiol group e.g.-SH.
The free valence indicated in each of the groups given in the preceding paragraph preferably attaches the nucleophilic centre to the carrier via a linker structure comprising the above- mentioned bivalent alkylene group -R**-. A hydrogen bound directly to nitrogen may be replaced with a monovalent alkyl group selected amongst the same alkyl groups as R*- as long as they are not substantially counteracting the desired reactivity of the unsubstituted form of the nucleophilic centre. Thus the hydrogen in a thiol group and in a hydroxyl group cannot be replaced, for instance. Two replacing alkyl groups may form a cyclic structure together with atom to which they are attached, i.e. form a bivalent alkylene group e.g.
selected amongst the alternatives for the -R**- group.
The bivalent structures -R**- and -R"- discussed above comprises next to the carrier a linker structure which does not negatively affect the desired effect of the nucleophilic centre of the active structure. Such structures are not part of the invention and suitable such structures can be designed by the average-skilled person in the field.
In alternative active structures there may be a second nucleophilic centre which a) may be part of one of the organic groups, e.g. the R*- or the -R**- group mentioned above, and b) exhibit a first heteroatom N, O or S (= Y1) in the same manner as for the first nucleophilic centre. In principle this means that this second nucleophilic centre complies with the formula:
Y^-R^X-R mHn (formula III)
and the formula
-CH2(Y4)0<C=Y3)n»(Y2)m"- (formula IV) where m, n, m", n", o ", Y1, Y2, Y3, Y4, -R"- and -R' are selected in the same groups of variables as m, n, m', η', o ', X1, X2, X3, X4, -R"- and -R' of formula I and II. This includes that hydrogens (H) may be replaced as suggested for formulae I and II.
That is, the second nucleophilic centre comprises a group selected amongst:
a) amino groups, preferably primary or secondary amino groups,
b) hydrazide groups such as -NH-NH2, e.g. as part of a -CONHNH2 group, a semicarbazide group such as -NHCONHNH2, a carbazate group such as -OCONHNH2, a thiosemicarbazide group such as -NHCSNHNH2, a thiocarbazate group such as -OCSNHNH2,
c) aminooxy groups, such as -ONH2 etc, and
d) thiol groups, e.g.-SH. The heteroatom Y1 preferably is part of
a) an -NH2 group where the free valence preferably may bind to a sp3-hybridised carbon, or b) a thiol group -SH where the free valence preferably may bind to a sp3-hybridised carbon. Each of m", n" and o" in formula IV is 0 in both (a) and (b).
The distance between the first heteroatom Y1 and the first heteroatom X1 is typically larger than two or three atoms with upper limits being e.g. 20 atoms with preference for 4, 5 or 6 atoms between these two heteroatoms. The distance should support intra-molecular cyclisation, typically via one or more addition reactions.
The carrier
The selection of suitable carriers depends on the requirements of a particular use. The typical carrier is selected amongst macro molecular compounds, i.e. is a compound which has a molecular weight of > 2000 dalton, preferably > 10000 dalton or > 50000 dalton, and preferably exhibits a polymeric structure, i.e. is a polymer which may be a homopolymer, copolymer or a chemical adduct between two or more polymers of different polymeric structure. Other suitable carriers may have molecular weights < 2000 dalton and
exhibitpolymeric structure as indicated by the possibility of the low numbers of monomeric units discussed below, e.g. > 20 and < 100. The term "adduct polymer" in this context means a product formed by reacting two polymers exhibiting mutually reactive groups capable of forming covalent bonds that link the two polymers together upon reaction of the two mutually reactive groups with each other. See for instance WO 2009108100 (IPR-Systems AB) and references cited therein. Suitable macromolecular carriers may thus be selected amongst synthetic polymers (= man-made polymers), biopolymers (nature-made polymers such as polysaccharides, polypeptides, proteins etc) and biosynthetic polymers where "biosynthetic polymer" refers to a macromolecular carrier or compound exhibiting both a synthetic polymeric structure and a biopolymeric structure. A carrier polymer may be cross- linked or not cross-linked. With respect to branching the polymer may be unbranched, i.e. linear, or branched including either hyperbranched or dendritic. The degree of branching may thus vary between 0 and 1, such as be > 0.10 or > 0.25 > 0.5 > 0.75 or > 0.90 and/or < 0.90 or < 0.75 or < 0.50 or < 0.25 or < 0.10. Cross-linked polymers are as a rule insoluble in aqueous liquids while the solubility of non-cross-linked polymers depend on the overall structure of the polymer, e.g. presence and amount of polar and/or hydrophilic groups. Carrier polymers may also be derivatized to contain non-polymeric or polymeric groups, for instance cross-links, substituents, charged or uncharged groups, active structures (as discussed above) etc. Macromolecular carriers which are insoluble in aqueous liquids may have different physical and geometric shapes as discussed for support materials elsewhere in this specification.
The term polymer above includes organic as well as inorganic polymers.
The macro molecule or polymer used in the carrier may be water-insoluble and suspended in aqueous liquid media (when in particle form).
Polymers and other macromolecules suitable as carrier material may be hydrophilic or hydrophobic with preference for hydrophilic. The introduction of hydrophilic groups may among others be accomplished by:
a) coating with a hydrophilic material,
b) selecting building blocks/monomers which exhibit hydrophilic groups and appropriate conditions during synthesis of the macromolecular compound, and
c) chemical derivatisation with hydrophilic groups subsequent to the synthesis of the basic hydrophobic polymer etc.
The hydrophilicity of a group, structure or carrier molecule increases as a rule with an increase in the ratio r = the sum of the number of heteroatoms O, N and S divided by the sum of the number of carbon atoms. Hydrophilic groups/compounds typically have an r > 0.5, preferably > 1.0, and for hydrophobic groups r < 1.0, preferably < 0.5. Typical hydrophilic groups are hydroxy, amino, amido, carboxy (including free acid carboxyl as well as carboxylate (ester ester and salt) etc. Typical hydrophobic groups are alkyls (CnH(2n+i)-, CnH(2n_i)-, CnH(2n_3)- etc), phenyls including alkyl phenyls, benzyl including other phenylalkyls etc. A carrier macromolecule typically comprises a polymer backbone which comprises > 5, or more preferably > 10 such as > 25 different and/or identical monomeric units linked together. The polymer may carry projecting or pending polymeric and/or non-polymeric groups of various lengths and kinds. A carrier polymer is preferably hydrophilic with hydrophilic groups selected amongst those given elsewhere in this specification. The most preferred hydrophilic group is hydroxy with the preferred carrier polymers and/or other macro molecular carrier being selected by poly hydroxy polymers (PHP or PH-polymers) exhibiting > 5, with preference for > 10, such as > 25 or > 50 hydroxyl groups and/or > 5 monomeric subunits each of which exhibits one, two, three, four or more hydroxyl groups per unit.
Typical polymers that may be present in polymeric carriers are a) polyester polymers, b) polyamide polymers, c) polyether polymers, d) polyvinyl polymers, e) polysaccharides etc. A carrier may comprise one or more of these polymers/polymeric structures.
Polyester polymers are in particular obtained by polymerisation of a) monomers exhibiting at least one hydroxy group and at least one carboxy group, or b) a mixture containing monomers exhibiting two or more hydroxy groups and monomers exhibiting two or more carboxy group.
Polyamide polymers are in particular obtained by polymerisation of a) monomers exhibiting at least one amino group and at least one carboxy group, or b) a mixture containing monomers exhibiting two or more amino groups and monomers exhibiting two or more carboxy group.
An important group of polyamides are those that exhibit polypeptide structure together with a plurality of hydroxy groups (PH-polymers). Suitable polyamide polymers of this kind are typically based on hydroxy-,amino-carboxylic acids as monomers, in particular with the amino group positioned a to the carboxylic group, e.g. serine, threonine, tyrosine, proline etc.
Polyether polymers are typically used in combination with other polymeric structures, e.g. polymers of (a), (b), (d) and/or (e) above, which are polyfunctional with respect to the presence of groups such as hydroxy, amino etc. Typical polyether polymers are polyethylene oxide and various copolymerisates between ethylene oxide and other lower alkylene oxides, lower epihalohydrins etc. Polyvinyl polymers which may be suitable as polymeric carriers in the invention are typically found amongst polymers containing one, two or more different monomeric units selected amongst hydroxyalkyl acrylates and methacrylates, N-hydroxyalkyl acryl- and N- hydroxyalkyl methacrylamides, hydroxyalkyl vinyl ethers, vinyl esters etc. In one
5 embodiment the carrier is a polyvinyl alcohol (PVA). Polyvinyl alcohols are typically obtained by partial hydrolysis of polyvinyl esters meaning that polyvinyl alcohols that are preferred in the invention typically exhibit residual amounts of ester groups (< 10 % or < 5%).
10 Typical polysaccharides that may be present in carriers used in the invention include dextran, starch, agarose, agaropektin, cellulose, glucosamino glucanes (GAG), and derivates of these polysaccharides etc. The most interesting polysaccharides are dextran, certain glucosamino glucanes (GAG) such as hyaluronic acid etc.
15 A polymer to be used in the carrier may have been derivatized, e.g. cross-linked and/or functionalized after its synthesis.
The active structure including the first, the optional second nucleophilic centre and the various heteroatoms discussed for the active structures are typically part of one and the same 20 organic group/substituent attached to the macromolecular carrier. In certain variants different parts of an active structure may be part of different groups/substituents attached to the carrier and/or part of the carrier.
Sizes/molecular weights of suitable carrier polymers will among others depend on the actual 25 application/use of the composition/method of the invention. Thus suitable polymeric carriers with respect to a particular polymeric structure and/or size may vary within a wide interval. Thus as a rule the number of monomeric subunits (mean value) of a polymer present in the carrier may be > 20 or > 100 or > 200 or > 300 or > 500 or > 1000 or > 2000 or > 20 000 or > 50 000 and/or < 50 000 or < 20 000 or < 2000 or < 1000 or < 500 or < 300 or < 200, or < 30 100 (with the proviso that >-limit always is lower than a <- limit when these values are
combined to define intervals). Preferred numbers of monomeric units may in some cases be found in the interval of 200 - 600 which in particular applies to the polyvinylalcohol used in the experimental part. Suitable numbers of active structures or nucleophilic centres per monomeric unit of a polymer of the carrier will also depend on the use, the active structure, etc and may thus be found within a wide interval, such as < 80 %, such as < 50% or < 20 % with typical lower limits being 0.01 % or 0.1 % or 1 % where 100% corresponds to one active structure or nucleophilic centre per monomeric unit. For active structures containing two or more nucleophilic centres the number of nucleophilic centres per monomeric unit may exceed 100 %, such as > 100% or > 125 % or > 150%. Other features of the composition
The AP is present in the composition as an AP-formulation in which the AP is:
a) in dry form, for instance as free particles,
b) in dissolved form, typically in an aqueous liquid medium, and
c) in suspended/dispersed form, i.e. as water-insoluble particles suspended in an aqueous liquid medium,
d) attached to a support which is insoluble in aqueous liquid media.
The term "dissolved" in this context means that the AP is present as a solute. The AP particles comprise the AP in a pure form or diluted with some solid material. Useful concentrations of the AP in formulations according to (b) can be found within a broad interval.
The composition may in addition to the AP contain an aqueous solution such as a buffer (e.g. PBS buffer), saline solution, salts etc. However the active principle may more or less be the only component of the composition.
Synthesis of the active principle
The AP may be synthesized according to well-known protocols, for instance of the kinds given in WO 2009108100 (IPR-Systems AB) and references cited therein. Water-insoluble support
As mentioned above the AP may be fixed to a water-insoluble support. This support material may be selected amongst support materials that have at least one or more of the following characteristics: a) in the form of particles, b) porous or non-porous particles or monoliths allowing or not allowing, respectively, aqueous liquids to penetrate the support, c) rigid, d) soft, e) elastic, f) compressible, g) gellable (in particular to form a hydrogel when placed in contact with water) etc. The support may comprise plastics, glass, mineral, metal etc. When the carrier of the AP is insoluble in aqueous media the carrier as such may define its own 5 solid support.
Suitable supports typically comprise polymeric materials, e.g. comprising one or more polymer selected from the same polymers as the carrier polymers are selected. Typical carrier polymers are polysaccharides, e.g. cellulose, cross-linked dextran, agarose such as 10 cross-linked agarose etc, polyester polymers e.g. lactic acid copolymers such as polyglactin, polyethylenes etc. In one embodiment the polymer is polyvinyl chloride (PVC). Other kinds of support material may also be used, e.g. ceramic materials, plastics, mineral materials, metals, composite material, activated carbon etc. Porous forms of theses materials may be used as filters and/or adsorbent material.
15
Attachment to the support may be accomplished by mixing, coating, impregnating etc the support with AP according to techniques known in the field. Alternatively, the
macro molecular carrier of AP may be part of the material from which a support/device is made.
20
Contacting AP with erythrocytes
As discussed above the contacting of the AP with the erythrocytes may take place in vivo of or separate from the individual to be treated.
25 The amount of the AP in the composition which is brought into contact with the red blood cells (erythrocytes) is effective in the sense that the cells are stabilized or preserved with reduced haemolysis as a result. The suitable dosage (per administration) for in-vivo applications, depends partly on formulation (e.g. the kind of support material, AP, , concentrations etc) etc and thus is selected within a broad interval, e.g. 5μg/ml or more, or
30 10μg/ml or more, or 20μg/ml or more, or 100μg/ml or more, or 200μg/ml or more, or
500μg/ml or more, or 20 mg/ml or less, but preferably 10 mg/ml or less, or 5 mg/ml or less, or 1 mg/ml or less. In one embodiment the concentration is 10μg/ml to lOmg/ml such as 100μg/ml to 1 mg/ml. Experimental testing is needed for individual cases. Typical devices to which the present invention can apply.
Containers suitable for containing erythrocytes may contain the AP. The container may be a bag, such as a blood bag, a syringe, a vial, a beaker, an e where the container is coated with the AP according to the invention. The container may be made of any suitable material such as a plastic material for example polyvinyl chloride (PVC).
The composition may be applied to the surface of the container by dip, spin or spray coating the surface of the container or using any other suitable technique. The composition may comprise a suitable solvent. Adding a solvent to the composition may make it easier to apply the composition to the surface of the container. The solvent may be an aqueous solvent such as water or a buffer (e.g. PBS buffer), or an alcohol such as ethanol.
A tube such as a dialysis tube may also be coated with the composition or AP. The inner surface of the tube facing the erythrocytes may be coated with the AP by applying the composition using any suitable technique as described above.
The surface of the support or the device (container or tube) may be pre-treated in order to enhance the attachment or adsorption of the composition or AP.
The amount of AP on the surface should be such that the stabilizing or preserving effect of the AP is obtained.
Other conceivable products are strips, chips, glass slides for blood testing or sampling or blood analysis where at least a part of its surface is coated with the composition or AP.
A mixture of stabilized erythrocytes
The present invention also relates to a mixture of erythrocytes and the composition according to the present invention. The erythrocytes are stabilized or preserved in the mixture and may therefore be stored for a longer time without extensive haemolysis. The mix may comprise 5μg/ml or more, or 10μg/ml or more, or 20μg/ml or more, or 100μg/ml or more, or 200μg/ml or more, or 500μg/ml or more, or 20 mg/ml or less, but preferably 10 mg/ml or less, or 5 mg/ml or less, or 1 mg/ml or less. EXPERIMENTAL PART
EXAMPLE 1 - SYNTHESIS OF CARBAZATE-FUNCTIONALIZED POLYVINYL ALCOHOL (PVAC) Polyvinyl alcohol (5 g, 13-23 kDa) was dissolved in dimethyl sulfoxide (100 mL) while stirring for 1 hour at 80°C under argon gas. Carbonyl diimidazole (10 g) was added and stirring continued at room temperature overnight. Hydrazine hydrate (10 mL) was then added, the reaction stirred overnight, and the product collected and purified by repeated precipitation in ethanol. The degree of substitution was determined spectrophotometrically by performing a trinitrobenzene sulfonic acid assay described elsewhere (Stephen L. Snyder and Philip Z. Sobocinski; Analytical Biochemistry 64, 284-288, 1975).
EXAMPLE 2 - STABILISATION OF ERYTHROCYTES Storage of red blood cells in DEHP-free conditions
In an attempt to replace DEHP in the storage of red blood cells PVAC dissolved in PBS buffer was added to the storage container. The concentration of PVAC in the blood was 20 μg/mL or 200 μg/mL. Hemolysis was measured throughout the storage period and demonstrated a dose-dependent decrease with the addition of PVAC (n = 3, * denotes statistically significant difference with p < 0.05). Pure PBS was used as control (K). The results are shown in Figure 1.

Claims

C L A I M S
1. A composition for use in preserving or stabilizing erythrocytes wherein the
composition comprises as an active principle (= AP) comprising a carrier which exhibits a plurality of active structureactive structures,
said active structureactive structures comprising a nucleophilic centre complying with the formula
X^-fT-X-fOmHn (formula I)
where
a) X1 is a single-bonded heteroatom selected amongst N, O and S and exhibits a free electron pair,
b) m is 0 or 1 and n is 1 or 2 with the sum of m plus n being 2 for X1 = N and 1 for X1 = S and O,
c) -R"- is a bivalent organic group providing attachment to the carrier via one of its free valences and direct attachment to the heteroatom X1 at the other one of its free valences, and
d) R'- is a monovalent organic group directly attached to the heteroatom X1 via its free valence.
2. The composition of claim 1, characterized in that either one or both of the organic groups R'- and -R"-, with preference for -R"- , comprise a structure of the formula:
-CH2(X4V(C=X3)n>(X2)m>- (formula II) where
a) each of m', n' and o' is 0 or 1, with preference for m' being 1 with further
preference for either one or both of n' and o' also being 1,
b) each of X2 , X3, and X4, is selected amongst NH and a heteroatom S or O, with preference for either one or both of X2 and X4 being selected amongst NH and O with further preference for X3 being selected amongst NH, O and S,
c) the left free valence provides binding to a monovalent alkyl group R*- or to the carrier via at least a bivalent alkylene group -R**-, each of which two groups comprises the methylene group -CH2- shown in formula II, and
d) the right free valence binds directly to the first heteroatom X1. The composition according to any one of claims 1-2, characterized in that the nucleophilic center comprises a group selected amongst
a) amino groups, preferably primary or secondary amino groups,
b) hydrazide groups such as -NH-NH2, e.g. as part of a -CONHNH2 group, a
semicarbazide group such as -NHCONHNH2, a carbazate group such as - OCONHNH2, a thiosemicarbazide group such as -NHCSNHNH2, a thiocarbazate group such as -OCSNHNH2,
c) aminooxy groups, such as -ONH2 etc, and
d) thiol groups, e.g.-SH.
The composition according to any one of claims 1-3, characterized in that the scavenger structure comprises a second nucleophilic centre which exhibits a single- bonded first heteroatom N, O or S (=Y!) and comprises a group selected amongst a) amino groups, preferably primary or secondary amino groups,
b) hydrazide groups such as -NH-NH2, e.g. as part of a -CONHNH2 group, a
semicarbazide group such as -NHCONHNH2, a carbazate group such as - OCONHNH2, a thiosemicarbazide group such as -NHCSNHNH2, a thiocarbazate group such as -OCSNHNH2,
c) aminooxy groups, such as -ONH2 etc, and
d) thiol groups, e.g.-SH.
The composition according to any one of claims 1-4, characterized in that the carrier is a macromolecular carrier and/or is water-soluble or water-insoluble and preferably exhibits polymer structure.
The composition according to any one of claims 1-5, characterized in that a) the carrier is water-insoluble and defines a support, and/or b) the active principle is attached to a water-insoluble support. The composition according to claim 1 wherein the active principle is carbazate- functionalized polyvinyl alcohol.
The composition according to claim 1 wherein the composition further comprises buffer or a saline solution.
9. A method for stabilizing or preserving erythrocytes characterized in that the method comprises the steps of:
i) providing a composition according to any one of claims 1 to 8, and
ii) contacting the composition with erythrocytes.
10. The method according to claim 9, characterized in that said composition is present at a concentration of 5μg/ml or more, or 10μg/ml or more, or 20μg/ml or more, or 100μg/ml or more, or 200μg/ml or more, or 500μg/ml or more, or 20 mg/ml or less, but preferably 10 mg/ml or less, or 5 mg/ml or less, or 1 mg/ml or less.
11. A mixture comprising erythrocytes and the composition according to any one of claims 1 to 8.
12. The mixture according to claim 11 wherein the concentration of the composition is 5μg/ml or more, or 10μg/ml or more, or 20μg/ml or more, or 100μg/ml or more, or 200μg/ml or more, or 500μg/ml or more, or 20 mg/ml or less, but preferably 10 mg/ml or less, or 5 mg/ml or less, or 1 mg/ml or less.
13. A container having an inner and an outer surface wherein the inner surface is coated with the composition or active principle according to any one of claims 1 to 8.
14. The container according to claim 13 wherein the container is a blood bag.
15. A dialysis tube having an inner and an outer surface wherein the inner surface is coated with the composition or active principle according to any one of claims 1 to 8.
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WO2021233534A1 (en) * 2020-05-20 2021-11-25 Pvac Medical Technologies Ltd Use of substance and pharmaceutical composition thereof, and medical treatments or uses thereof
WO2022191759A1 (en) 2021-03-09 2022-09-15 Cremed AB (publ) A polymer-based carbonyl scavenger for detection and treatment of ischemic injuries

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