EP1404379A2 - Procede d'inactivation virale utilisant un antioxydant - Google Patents

Procede d'inactivation virale utilisant un antioxydant

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Publication number
EP1404379A2
EP1404379A2 EP02744213A EP02744213A EP1404379A2 EP 1404379 A2 EP1404379 A2 EP 1404379A2 EP 02744213 A EP02744213 A EP 02744213A EP 02744213 A EP02744213 A EP 02744213A EP 1404379 A2 EP1404379 A2 EP 1404379A2
Authority
EP
European Patent Office
Prior art keywords
quencher
blood
solution
additive solution
photosensitizer
Prior art date
Legal status (The legal status 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 status listed.)
Withdrawn
Application number
EP02744213A
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German (de)
English (en)
Inventor
Raymond P. Goodrich
Heather Reddy
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Terumo BCT Inc
Original Assignee
Gambro Inc
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Filing date
Publication date
Application filed by Gambro Inc filed Critical Gambro Inc
Publication of EP1404379A2 publication Critical patent/EP1404379A2/fr
Withdrawn legal-status Critical Current

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2/00—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor
    • A61L2/0005—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts
    • A61L2/0011—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts using physical methods
    • A61L2/0029—Radiation
    • A61L2/0052—Visible light
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K41/00—Medicinal preparations obtained by treating materials with wave energy or particle radiation ; Therapies using these preparations
    • A61K41/10—Inactivation or decontamination of a medicinal preparation prior to administration to an animal or a person
    • A61K41/17—Inactivation or decontamination of a medicinal preparation prior to administration to an animal or a person by ultraviolet [UV] or infrared [IR] light, X-rays or gamma rays
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2/00—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor
    • A61L2/0005—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts
    • A61L2/0011—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts using physical methods
    • A61L2/0029—Radiation
    • A61L2/0058—Infrared radiation
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2/00—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor
    • A61L2/0005—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts
    • A61L2/0011—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts using physical methods
    • A61L2/0029—Radiation
    • A61L2/0076—Radiation using a photocatalyst or photosensitiser
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2/00—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor
    • A61L2/0005—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts
    • A61L2/0082—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts using chemical substances
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2/00—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor
    • A61L2/0005—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts
    • A61L2/0082—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts using chemical substances
    • A61L2/0088—Liquid substances
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2/00—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor
    • A61L2/02—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor using physical phenomena
    • A61L2/08—Radiation
    • A61L2/10—Ultraviolet radiation
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61M—DEVICES FOR INTRODUCING MEDIA INTO, OR ONTO, THE BODY; DEVICES FOR TRANSDUCING BODY MEDIA OR FOR TAKING MEDIA FROM THE BODY; DEVICES FOR PRODUCING OR ENDING SLEEP OR STUPOR
    • A61M1/00—Suction or pumping devices for medical purposes; Devices for carrying-off, for treatment of, or for carrying-over, body-liquids; Drainage systems
    • A61M1/36—Other treatment of blood in a by-pass of the natural circulatory system, e.g. temperature adaptation, irradiation ; Extra-corporeal blood circuits
    • A61M1/3681—Other treatment of blood in a by-pass of the natural circulatory system, e.g. temperature adaptation, irradiation ; Extra-corporeal blood circuits by irradiation
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61M—DEVICES FOR INTRODUCING MEDIA INTO, OR ONTO, THE BODY; DEVICES FOR TRANSDUCING BODY MEDIA OR FOR TAKING MEDIA FROM THE BODY; DEVICES FOR PRODUCING OR ENDING SLEEP OR STUPOR
    • A61M1/00—Suction or pumping devices for medical purposes; Devices for carrying-off, for treatment of, or for carrying-over, body-liquids; Drainage systems
    • A61M1/36—Other treatment of blood in a by-pass of the natural circulatory system, e.g. temperature adaptation, irradiation ; Extra-corporeal blood circuits
    • A61M1/3681—Other treatment of blood in a by-pass of the natural circulatory system, e.g. temperature adaptation, irradiation ; Extra-corporeal blood circuits by irradiation
    • A61M1/3683—Other treatment of blood in a by-pass of the natural circulatory system, e.g. temperature adaptation, irradiation ; Extra-corporeal blood circuits by irradiation using photoactive agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2202/00—Aspects relating to methods or apparatus for disinfecting or sterilising materials or objects
    • A61L2202/20—Targets to be treated
    • A61L2202/22—Blood or products thereof

Definitions

  • photosensitizers that have been proposed for use for blood component photoirradiation have undesirable properties.
  • European Patent Application 196,515 published Oct. 8, 1986 suggests the use of non-endogenous photosensitizers such as porphyrins, psoralens, acridine, toluidines, flavine (acriflavine hydrochloride), phenothiazine derivatives, and dyes such as neutral red and methylene blue, as blood additives.
  • Another molecule, chlorpromazine has been used as a photosensitizer; however its usefulness is limited by the fact that it should be removed from any fluid administered to a patient after the decontamination procedure because it has a sedative effect.
  • Protoporphyrin which occurs naturally within the body, can be metabolized to form a photosensitizer; however, its usefulness is limited in that it degrades the desired biological activities of proteins.
  • Pathogen kill using riboflavin and related photosensitizer compounds occurs upon photoinactivation via oxidative damage, including singlet oxygen damage, or via binding of the photosensitizer to nucleic acids of the pathogen, thereby disrupting the ability of the pathogen to function and reproduce, or both. Side reactions of photolysis, may cause damage to blood products and compromise their suitability for transfusions.
  • the invention is directed to a method for quenching side reactions generated by a photosensitizer and light.
  • the side reactions may cause unwanted damage to desired biological components.
  • the invention is more specifically directed to a method of inactivating pathogens in a blood component which may contain pathogens comprising: adding a photosensitizer and a quencher to the blood component and illuminating the blood component, photosensitizer and quencher for a time sufficient to inactivate any pathogens contained therein.
  • the quencher may also be added after illumination of the blood component and photosensitizer.
  • the quencher may also be added to a storage solution used to store the blood or blood component after irradiation.
  • a dry composition is provided which is adapted to be mixed with a solvent, comprising a quencher and a photosensitizer.
  • the invention is further directed towards an additive solution comprising at least one quencher for quenching side reactions generated in a solution containing blood or blood component and a photosensitizer by application of light energy.
  • This invention also provides a fluid comprising biologically active protein, blood or blood constituents and inactivated microorganisms, endogenous photosensitizer or photoproduct thereof, and one or more quenchers.
  • this invention is directed towards reducing damage to cells from side reactions that occur when a composition containing red blood cells and riboflavin is irradiated with visible light without interfering with the inactivation of pathogens by riboflavin.
  • blocking agent or “quenching agent” or “quencher” refer to compounds that reduce damage to biological components caused by the compounds and/or conditions used for pathogen inactivation.
  • Some side reactions which may damage cell membrances of blood and blood components include formation of reactive oxygen species from the application of visible light to solutions containing photosensitizers. These side reactions are known to one of ordinary skill in the art.
  • the specific use of a quencher can selectively protect cell membranes from oxidation damage while permitting oxygen-based chemistry to proceed in solution where virus and other pathogenic agents primarily reside. In this way, the positive effects of oxidation chemistry can be utilized for pathogen kill while the negative side-effects of cell and protein damage are eliminated.
  • quenching agents are thiol containing compounds including glutathione, n-acetyl-cysteine, and cysteine.
  • Another class of quenching agents are amino acids such as tryptophan, tyrosine, histidine, adenine and methionine.
  • Another class of quenching agents are antioxidants, including Vitamin E, TPGS, trolox and Vitamin C and their derivatives.
  • Antioxidants as well as thiol-containing compounds may help prevent denaturation of proteins in blood by side reactions during the irradiation process.
  • Amino acids may act as false targets for the side reactions which occur during the irradiation process, thus protecting the blood proteins.
  • Quenchers may also be added to the fluid to make the process more efficient and selective.
  • quenchers include antioxidants or other agents to prevent damage to desired fluid components or to improve the rate of inactivation of microorganisms and are exemplified by adenine, histidine, cysteine, tyrosine, tryptophan, ascorbate, N-acetyl-L- cysteine, propyl gallate, glutathione, mercaptopropionylglycine, dithiothreotol, nicotinamide, BHT, BHA, lysine, serine, methionine, glucose, mannitol, trolox, glycerol, vitamin E and mixtures thereof.
  • Quenchers can be used in any effective amount. All specific concentration amounts and intermediate ranges are included in the disclosure. A preferred range is from 0.05 to about 100 millimolar, more preferably from 0.1 to 20 millimolar. When used as a quencher, thiol containing compounds are preferably added in an amount of between 0.5 - 50 mM, and most preferably between about 0.1 - 20 mM. When used as a quencher, Vitamin E is preferably added in an amount of between about 0.1 and about 200 IU/ml, and more preferably about 0.1 to about 1 IU/ml.
  • Vitamin C When used as a quencher, Vitamin C is preferably added in an amount of between about 0.1 to 100 millimolar, preferably about 5 to about 15 millimolar, and more preferably about 10 millimolar.
  • concentrations of other quenchers may be determined by one of ordinary skill in the art without undue experimentation by analysis of the desired level of microorganism inactivation, the desired photosensitizer and other parameters, as taught herein and known to one of ordinary skill in the art.
  • quenching agents can be used in a variety of different ways in the methods of the invention. Quenchers may be added to the blood component at any stage of the process. For example, the quenchers may be added before adding photosensitizer and exposure to light, or may be added after exposure of the blood component to light. Quenching agents can be added to a solution used to store blood or blood components. Quenching agents can be added to blood or blood components along with the desired photosensitizer. Compositions containing quenching agents can be used in a dry form or a liquid form. Some preferred uses are given below. The quenchers of the invention may be used with platelet and red blood cell additive solutions and other blood component additive solutions as known in the art.
  • the quencher react with the pathogen inactivating compound to quench side reactions without interfering with pathogen inactivation by the pathogen inactivating compound. This may occur, for example, by the use of a quencher which substantially does not traverse pathogen membranes, and therefore quenches side reactions outside the pathogen membranes. In another embodiment, the quencher may react kinetically so slowly with the pathogen inactivating compound, or reactive intermediates formed therefrom, that it substantially does not interfere with pathogen inactivation. It is also preferred that the quencher does not negatively interact with the blood or blood components to be pathogen inactivated.
  • An effective amount of quencher is an amount that reduces undesired side reactions generated by illumination of a photosensitizer without causing the effectiveness of the photosensitizer to inactivate microorganisms to fall below a desired level.
  • the method of this invention improves the quality of blood or blood components treated with a pathogen-inactivation procedure, permitting more stringent pathogen inactivation processes to be utilized without compromising the integrity of the therapeutic components.
  • An amount of photosensitizer higher than that used in previously- known methods for inactivation of microorganisms may be used when a quencher is added to offset damage to desired components of the fluid from photoirradiation.
  • Decontamination methods of this invention using quenchers, endogenous photosensitizers and endogenously-based photosensitizer derivatives do not substantially destroy the biological activity of fluid components other than microorganisms. As much biological activity of these components as possible is retained, although in certain instances, when the methods are optimized, some loss of biological activity, e.g., denaturization of protein components, must be balanced against effective decontamination of the fluid. So long as fluid components retain sufficient biological activity to be useful for their intended or natural purposes, their biological activities are not considered to be "substantially destroyed.”
  • the quenchers, endogenous photosensitizers and endogenously-based derivative photosensitizers disclosed herein can be used in pre-existing blood component decontamination systems as well as in the decontamination system disclosed herein.
  • the endogenous photosensitizers and endogenously-based derivative photosensitizers of this invention can be used in the decontamination systems described in U.S. Pate ⁇ . Nos. 5,290,221, 5,536,238, 5,290,221 and 5,536,238.
  • Figure 1 depicts inactivation of bacteria as a function of platelet preparation and energy of irradiation, using 90% platelets in plasma and 10% platelet additive solution (90: 10) and 30% platelets with 70% additive solution (30:70).
  • Figure 2 shows the effect on inactivation of virus, bacteriophage and bacteria of adding antioxidants to platelet concentrate.
  • Figure 3 shows the inactivation curve for Herpes Simplex type II virus as a function of concentration of photosensitizer at an energy of irradiation of 20J/cm 2 using half ultraviolet and half visible light.
  • Figure 4 shows inactivation of S. epidermidis at varying concentrations of photosensitizer and energies of irradiation.
  • Figure 5 shows inactivation of ⁇ XI 74 at varying concentrations of photosensitizer and energies of irradiation.
  • Figure 6 shows inactivation of S. aureus and ⁇ XI 74 at varying energies of irradiation using a 50:50 mixture of ultraviolet and visible light.
  • Figure 7 shows inactivation of S. epidermidis and HSV-II at varying energies of irradiation using a 50:50 mixture of ultraviolet and visible light.
  • Figure 8 shows inactivation of HSV2 virus in blood bags agitated and irradiated at varying energy levels.
  • Figure 9 compares inactivation results for vaccinia virus in various fluids using ultraviolet light alone or 50:50 visible and ultraviolet light.
  • Figure 10 compares inactivation results with and without sensitizer of vaccinia virus at varying irradiation times.
  • Figure 11 compares inactivation of extracellular H1V-1 at 5 and 50 ⁇ M of photosensitizer and varying irradiation energies.
  • Figure 12 compares inactivation of intracellular HIV-l at 5 and 50 ⁇ M of photosensitizer and varying irradiation energies.
  • Figure 13 compares inactivation of intracellular HJN-1 at 5 and 50 ⁇ M of photosensitizer and varying irradiation energies, using p24 antigen levels.
  • Figure 14 shows inactivation of HSN-II at varying irradiation levels using platelet concentrate and platelet concentrate in media containing platelet additive solution with ascorbate.
  • Figure 15 shows an embodiment of this invention using a blood bag to contain the fluid being treated and photosensitizer and a shaker table to agitate the fluid while exposing to photoradiation from a light source.
  • Figure 16 shows an embodiment of this invention using blood bags which are prepackaged to contain the photosensitizer necessary for inactivation of contaminants in the blood or other bodily fluid.
  • Figure 17 shows an embodiment of this invention using blood bags as in Figure 16 with a container in the tubing line between the bags.
  • Figure 18 shows BNDV kill in red blood cells using different treatment conditions over time.
  • Figure 19 compares the log reduction of BVDV in washed red blood cells as compared to un-washed red blood cells with and without quencher.
  • Figure 20 shows the inactivation kinetics (log virus titre) of BVDV virus in the presence and absence of Vitamin E in platelets.
  • Figure 21 shows the level of inactivation observed between samples with and without Vitamin E in platelets.
  • Figure 22 shows the survival of radiolabelled primate platelets under varying treatment conditions (with and without antioxidants and untreated controls).
  • a "photosensitizer” is defined as any compound which absorbs radiation of one or more defined wavelengths and subsequently utilizes the absorbed energy to carry out a chemical process.
  • Photosensitizers used in this invention may include compounds which preferentially adsorb to nucleic acids, thus focusing their photodynamic effect upon microorganisms and viruses with little or no effect upon accompanying cells or proteins.
  • Other photosensitizers are also useful in this invention, such as those using type I (free- radical) and/or type II (singlet oxygen) dependent mechanisms.
  • endogenous photosensitizers Most preferred are endogenous photosensitizers.
  • endogenous means naturally found in a human or mammalian body, either as a result of synthesis by the body or because of ingestion as an essential foodstuff (e.g. vitamins) or formation of metabolites and/or byproducts in vivo.
  • endogenous photosensitizers are alloxazines such as 7,8-dimethyl-10-ribityl isoalloxazine (riboflavin), 7,8,10-trimethylisoalloxazine (lumiflavin), 7,8-dimethylalloxazine (lumichrome), isoalloxazine-adenine dinucleotide (flavine adenine dinucleotide [FAD]), alloxazine mononucleotide (also known as flavine mononucleotide [FMN] and riboflavine-5 -phosphate), vitamin Ks, vitamin L, their metabolites and precursors, and napththoquinones, naphthalenes, naphthols and their derivatives having planar molecular conformations.
  • alloxazines such as 7,8-dimethyl-10-ribityl isoalloxazine (riboflavin), 7,8,10-trimethylisoalloxazine (lumif
  • alloxazine includes isoalloxazines.
  • Endogenously-based derivative photosensitizers include synthetically derived analogs and homologs of endogenous photosensitizers which may have or lack lower (1-5) alkyl or halogen substituents of the photosensitizers from which they are derived, and which preserve the function and substantial non-toxicity thereof.
  • endogenous photosensitizers particularly when such photosensitizers are not inherently toxic or do not yield toxic photoproducts after photoradiation, no removal or purification step is required after decontamination, and treated product can be directly returned to a patient's body or administered to a patient in need of its therapeutic effect.
  • Preferred endogenous photosensitizers are:
  • Pathogen inactivation requires mixing the photosensitizer with the material to be decontaminated. Mixing may be done by simply adding the photosensitizer or a solution containing the photosensitizer to the fluid to be decontaminated.
  • the material to be decontaminated to which the photosensitizer has been added is flowed past a photoradiation source, and the flow of the material generally provides sufficient turbulence to distribute the photosensitizer throughout the fluid to be decontaminated.
  • the fluid and photosensitizer are placed in a photopermeable container and irradiated in batch mode, preferably while agitating the container to fully distribute the photosensitizer and expose all the fluid to the radiation.
  • quenchers can be added to the system at any desired stage in the process including mixing with the photosensitizer prior to irradiation.
  • the amount of photosensitizer to be mixed with the fluid will be an amount sufficient to adequately inactivate microorganisms therein, but less than a toxic (to humans or other mammals) or insoluble amount.
  • optimal concentrations for desired photosensitizers may be readily determined by those skilled in the art without undue experimentation.
  • the photosensitizer is used in a concentration of at least about 1 ⁇ M up to the solubility of the photosensitizer in the fluid, and preferably about 10 ⁇ M.
  • concentration range between about 1 ⁇ M and about 500 ⁇ M is preferred, preferably about 50 ⁇ M for platelets and plasma and preferably about 450 ⁇ M for red blood cells.
  • the fluid to be treated is exposed to photoradiation of the appropriate wavelength to activate the photosensitizer, using an amount of photoradiation sufficient to activate the photosensitizer as described above, but less than that which would cause non-specific damage to the biological components or substantially interfere with biological activity of other proteins present in the fluid.
  • the wavelength used will depend on the photosensitizer selected, as is known in the art or readily determinable without undue experimentation following the teachings hereof.
  • U.S. patent 6,258,577 and continuation in part 6,277,337 hereby incorporated by reference to the extent not inconsistent with the disclosure herein, describes methods and apparatus for pathogen inactivation of biological contaminants using endogenous photosensitizers, including 7,8-dimethyl-lO-ribityl isoalloxazine (riboflavin).
  • U.S. Patent 6,268,120 hereby incorporated by reference to the extent not inconsistent with the disclosure herein also describes pathogen inactivation of biological contaminants using isoalloxazine derivatives. Inactivation methods using endogenous photosensitizers are also described in U.S. Patent Application Serial No.
  • Microorganisms include viruses (both extracellular and intracellular), bacteria, bacteriophages, fungi, blood-transmitted parasites, and protozoa.
  • Exemplary viruses include acquired immunodeficiency (HIN) virus, hepatitis A, B and C viruses, sinbis virus, cytomegalovirus, vesicular stomatitis virus, herpes simplex viruses, e.g. types I and II, human T-lymphotropic retroviruses, HTLV-HI, lymphadenopathy virus LAV/TDAV, parvovirus, transfusion-transmitted (TT) virus, Epstein-Barr virus, and others known to the art.
  • HIN acquired immunodeficiency
  • hepatitis A, B and C viruses sinbis virus
  • cytomegalovirus vesicular stomatitis virus
  • herpes simplex viruses e.g. types I and II, human T-lymphotropic retroviruses
  • HTLV-HI lymphaden
  • Bacteriophages include ⁇ X174, ⁇ 6, ⁇ , R17, T 4 , and T 2 .
  • Exemplary bacteria include P. aeruginosa, S. aureus, S. epidermis, L. monocytogenes, E. coli, K. pneumonia and S. marcescens.
  • Inactivation of white blood cells may be desirable when suppression of immune or autoimmune response is desired, e.g., in processes involving transfusion of red cells, platelets or plasma when donor white blood cells may be present.
  • Materials which may be treated by the methods of this invention include any materials which are adequately permeable to photoradiation to provide sufficient light to achieve viral inactivation, or which can be suspended or dissolved in fluids which have such permeability to photoradiation.
  • materials are whole blood and aqueous compositions containing biologically active proteins derived from blood or blood constituents. Packed red cells, platelets and plasma (fresh or fresh frozen plasma) are exemplary of such blood constituents.
  • therapeutic protein compositions containing proteins derived from blood such as fluids containing biologically active protein useful in the treatment of medical disorders, e.g.
  • factor Vm Von Willebrand factor
  • factor IX factor X
  • factor XI factor XI
  • Hageman factor prothrombin
  • anti-thrombin HI fibronectin
  • plasminogen plasma protein fraction
  • immune serum globulin modified immune globulin
  • albumin plasma growth hormone
  • somatomedin plasminogen streptokinase complex
  • ceruloplasmin transferrin
  • haptoglobin antitrypsin and prekallikrein
  • Other fluids which could benefit from the treatment of this invention are peritoneal solutions used for peritoneal dialysis which are sometimes contaminated during connection, leading to peritoneal infections.
  • the material to be treated is preferably a fluid that comprises blood constituents, e.g. whole blood, platelets, plasma, white blood cells, or red blood cells.
  • the fluid may be a separated blood product.
  • the method is especially useful for treating platelets.
  • Vitamin E as a quencher is preferred for platelet treatment.
  • thiol-containing compounds such as glutathione are preferably used as quenchers.
  • biologically active means capable of effecting a change in a living organism or component thereof.
  • biologically active with respect to “biologically active protein” as referred to herein does not refer to proteins which are part of the microorganisms being inactivated.
  • non-toxic with respect to the photosensitizers means low or no toxicity to humans and other mammals, and does not mean non-toxic to the microorganisms being inactivated.
  • Substantial destruction" of biological activity means at least as much destruction as is caused by porphyrin and porphyrin derivatives, metabolites and precursors which are known to have a damaging effect on biologically active proteins and cells of humans and mammals'.
  • substantially non-toxic means less toxic than porphyrin, porphyrin derivatives, metabolites and precursors that are known for blood sterilization.
  • blood product includes blood constituents and therapeutic protein compositions containing proteins derived from blood as defined above. Fluids containing biologically active proteins other than those derived from blood may also be treated by the methods of this invention.
  • a "solution useful for storing blood components" is any of a number of solutions that are known in the art to be useful for storing blood components. Examples are given herein. Any suitable container and apparatus may be used for irradiation, as known in the art. Either flow through or batch treatment may be used. In an embodiment involving batch-wise treatment, the fluid to be treated is placed in a photopermeable container which is agitated and exposed to photoradiation for a time sufficient to substantially inactivate the microorganisms.
  • the photopermeable container is preferably a blood bag made of transparent or semitransparent plastic, and the agitating means is preferably a shaker table.
  • the photosensitizer and quencher may be added to the container in dry form as a powder, tablet, capsule or pill or in liquid form and the container agitated to mix the photosensitizer and quencher with the fluid and to adequately expose all the fluid to the photoradiation to ensure inactivation of microorganisms.
  • Quencher and/or photosensitizer may be added to or flowed into the photopermeable container separately from the fluid being treated or may be added to the fluid prior to placing the fluid in the container.
  • photosensitizer and quencher are added to anticoagulant and the mixture of photosensitizer, quencher and anticoagulant are added to the fluid.
  • the photosensitizer and quencher may be added to the photopermeable container before sterilization of such container or after sterilization. Quenchers may be added in any effective amount.
  • a preferred range is from 0.05 to 100 millimolar, more preferably from 0.1 to 20 millimolar.
  • the photoradiation source may be connected to the photopermeable container for the fluid by means of a light guide such as a light channel or fiber optic tube which prevents scattering of the light between the source and the container for the fluid, and more importantly, prevents substantial heating of the fluid within the container.
  • a light guide such as a light channel or fiber optic tube which prevents scattering of the light between the source and the container for the fluid, and more importantly, prevents substantial heating of the fluid within the container.
  • Direct exposure to the light source may raise temperatures as much as 10 to 15 ° C, especially when the amount of fluid exposed to the light is small, which can cause denaturization of blood components.
  • Use of the light guide keeps any heating to less than about 2 ° C.
  • the method may also include the use of temperature sensors and cooling mechanisms where necessary to keep the temperature below temperatures at which desired proteins in the fluid are damaged.
  • the temperature is kept between about 0°C and about 45°C, more preferably between about 4°C and about 37°C, and most preferably about 22°C.
  • the photoradiation source may be a source of visible radiation or ultraviolet radiation or both.
  • the photoradiation source may be a simple lamp or may consist of multiple lamps radiating at differing wavelengths.
  • the photoradiation source should be capable of delivering from about 1 to at least about 120 J/cm 2 .
  • any means for adding the photosensitizer and quencher to the fluid to be decontaminated and for placing the fluid in the photopermeable container known to the art may be used, such means typically including flow conduits, ports, reservoirs, valves, and the like.
  • the system includes means such as pumps or adjustable valves for controlling the flow of the quencher and photosensitizer into the fluid to be decontaminated so that its concentration may be controlled at effective levels as described above.
  • photosensitizer and quencher are mixed with the anticoagulant feed to a blood apheresis system.
  • the pH of the solution is preferably kept low enough, as is known to the art, to prevent detachment of the sugar moiety.
  • the photosensitizer and/or quencher can be added to the fluid to be decontaminated in a pre-mixed aqueous solution, e.g., in water or storage buffer solution.
  • the photosensitizer and quencher are added to the fluid to be decontaminated as dry medium in powder, pill, tablet or capsule form.
  • the fluid is placed in a photopermeable container such as a blood bag, e.g. used with the apheresis system described in U.S. Patent No. 5,653,887, and agitated while exposing to photoradiation.
  • a photopermeable container such as a blood bag
  • Suitable bags include collection bags as described herein. Collection bags used in the SpectraTM system or TrimaTM apheresis system of Gambro BCT, Inc., of Lakewood, Colorado, are especially suitable. Shaker tables are known to the art, e.g. as described in U.S. Patent 4,880,788.
  • the bag is equipped with at least one port for adding fluid thereto.
  • the photosensitizer preferably 7,8-dimethyl-l 0- ribityl-isoalloxazine, and quencher, are added to the fluid-filled bag in dry form as a powder, pill, tablet or capsule.
  • the bag is then placed on a shaker table and agitated under photoradiation until substantially all the fluid has been exposed to the photoradiation.
  • the bag may be prepackaged with the powdered photosensitizer and quencher contained therein.
  • the fluid to be decontaminated may then be added through the appropriate port.
  • Decontamination systems as described above may be designed as stand-alone units or may be easily incorporated into existing apparatuses known to the art for separating or treating blood being withdrawn from or administered to a patient.
  • blood- handling apparatuses include the Gambro BCT, Inc., of Lakewood, Colorado, SpectraTM or
  • TRIMA ® apheresis systems available from Gambro BCT, Inc., of Lakewood, Colorado, or the apparatuses described in U.S. Patent 5,653,887 and U.S. Serial No. 08/924,519 filed September 5, 1997 (PCT Publication No. WO 99/11305) of Gambro BCT, Inc., of Lakewood, Colorado, as well as the apheresis systems of other manufacturers.
  • the decontamination system may be inserted just downstream of the point where blood is withdrawn from a patient or donor, just prior to insertion of blood product into a patient, or at any point before or after separation of blood constituents.
  • the photosensitizer and quencher are added to blood components along with anticoagulant in a preferred embodiment, and separate irradiation sources and cuvettes are placed downstream from collection points for platelets, for plasma and for red blood cells.
  • separate irradiation sources and cuvettes are placed downstream from collection points for platelets, for plasma and for red blood cells.
  • the use of three separate blood decontamination systems is preferred to placement of a single blood decontamination system upstream of the blood separation vessel of an apheresis system because the lower flow rates in the separate component lines allows greater ease of irradiation.
  • decontamination systems of this invention may be used to process previously collected and stored blood products.
  • the fluid may be thinned, exposed to higher energies of radiation for longer periods, agitated for longer periods or presented to photoradiation in shallower containers or conduits than necessary for use with other blood components.
  • the decontamination method of this invention using endogenous photosensitizers and endogenously-based derivative photosensitizers is exemplified herein using 7,8-dimefhyl-lO- ribityl isoalloxazine as the photosensitizer, however, any photosensitizer may be used which is capable of being activated by photoradiation to cause inactivation of microorganisms.
  • IgG is an immunoglobulin plasma protein that when present is caused to bind with the cells during the photoactivation process.
  • IgG The presence of IgG and the potential for it to become bound to the membrane of red cells raises potential concerns and difficulties in using photosensitizers to inactivate pathogens in red blood cells.
  • Physiological issues include immediate clearance of the treated red cells by the transfused patients' reticuloendothelial system and complement activation. Perhaps as important a concern with respect to IgG binding, even if the presence of IgG has no effect on cell survival or product safety, is with respect to the diagnostic implications.
  • the IgG bound to the treated cells cannot be removed from the cell membranes by washing the cells; even if extensive washings are performed. Because of the binding of the IgG to the cells, the cells may exhibit a positive test result when direct antiglobulin test (DAT or Coombs') is employed.
  • DAT direct antiglobulin test
  • the Coombs' test is used to detect antibody on red blood cells.
  • the test uses rabbit antisera to immunoglobulin. When cells coated with IgG are mixed with the rabbit antisera, agglutination occurs. If IgG coated red cells are transfused into a patient, a physician loses one of his important diagnostic tests in understanding hematologic changes in the patient, since patients receiving such a product may exhibit a positive Coombs' test.
  • quenchers help prevent IgG binding to red blood cells, and any consequent positive DAT results.
  • These quenchers may be added to the blood or blood components to be treated prior to, simultaneously with or after the addition of the pathogen inactivating compound as described above.
  • the quencher is added prior to or simultaneously with the pathogen inactivating compound.
  • the quenchers may be added to the biological material to be photoinactivated either alone or in combination with other quenchers.
  • quenchers which substantially do not damage blood components, and in particular do not damage red blood cell function or modify red blood cells after treatment. Also preferred are quenchers which do not substantially interfere with the pathogen inactivating compound.
  • the lack of a substantially damaging effect on red blood cell function may be measured by methods known in the art for testing red blood cell function. For example, the levels of indicators such as intracellular ATP (adenosine 5'- triphosphate), intracellular 2,3-DPG (2,3-diphosphoglycerol) or extracellular potassium may be measured, and compared to an untreated control. Additionally hemolysis, pH, hematocrit, hemoglobin, osmotic fragility, glucose consumption and lactate production may be measured.
  • concentrations of the pathogen inactivating compound and the quenching agent are given above, it is understood that the concentration of the quenching agent may be adjusted as needed in the blood product being treated to produce the desired reduction of unwanted side reactions, while still protecting the property of the material, such as red blood cell function, and also achieving the desired log kill of pathogens. These adjustments are known to one of ordinary skill in the art.
  • Some quenchers may oxidize or otherwise degrade or react over time.
  • the quencher when the quencher is a thiol containing compound, the quencher may oxidize to form disulfide dimers. It is preferred to add the quencher to the material at a time and concentration such that the quencher can quench the pathogen inactivating compound before it has substantially degraded or otherwise reacted in situ.
  • the addition of glutathione to the blood or blood component close to the time of addition of the pathogen inactivating compound is advantageous to minimize possible reduction of glutathione concentration, for example, by oxidation or peptidolysis, which may occur, for example in some biological materials, such as plasma.
  • the methods of the present invention are generally also useful to inactivate any biological contaminant found in stored blood or blood products, including bacteria and blood-transmitted parasites.
  • monoclonal or polyclonal antibodies directed against specific viral antigens may be covalently coupled with a photosensitizer compound.
  • Other fields of application wherein the present invention may find application include the preparation of non-infectious viral vaccines, therapeutic treatment of immune system disorders by photophoresis, elimination of viable nucleated cells such as leukocytes via the cytotoxicity of nucleic acid binding photosensitizers, and possible treatment for certain accessible cancers and tumors.
  • illumination solutions useful for irradiation of blood and blood components are provided.
  • a preferred illumination solution used in the methods of the invention for red blood cells contains 500 ⁇ M riboflavin, 0.9% sodium chloride and 4 mM glutathione.
  • blood or blood components are prepared as described herein or as known in the art.
  • the blood or blood components are combined with an illumination solution and irradiated, as described herein.
  • the irradiated red blood cells may be stored in a commonly used storage solution, for example, AS3 as described in published PCT application WO 01/94349 (incorporated by reference in its entirety to the amount not inconsistent herewith) to which additional quencher, preferably glutathione, may be added.
  • Glutathione has many properties that make it particularly useful as a quencher. It is normally present in all cell types. It is not believed substantially to be passively able to pass through the membranes, such as cell membranes or lipid coats, of bacteria and lipid- enveloped viruses. At pH 7, glutathione is charged and in the absence of active transport does not penetrate lipid bilayers to any significant extent.
  • the fluid to be decontaminated along with photosensitizer and quencher in bags which are photopermeable or at least sufficiently photopermeable to allow sufficient radiation to reach their contents to activate the photosensitizer.
  • Sufficient photosensitizer is added to each bag to provide inactivation, preferably to provide a photosensitizer concentration of at least about 10 ⁇ M, and the bag is agitated while irradiating, preferably at about 1 to about 120 J/cm for a period of between about 6 and about 36 minutes to ensure exposure of substantially all the fluid to radiation.
  • visible light is used.
  • the photosensitizer and quencher may be added in dry form as powder, or a pill, tablet or capsule.
  • the fluid to be decontaminated may contain additives or anticoagulant solutions and the blood product or blood components may be stored in such solutions. As described herein, quencher may be added to the system at any point.
  • the method preferably uses endogenous photosensitizers, including endogenous photosensitizers which function by interfering with nucleic acid replication.
  • 7,8-dimethyl-10- ribityl isoalloxazine is the preferred photosensitizer for use in this invention.
  • the chemistry believed to occur between 7,8-dimethyl-10-ribityl isoalloxazine and nucleic acids does not proceed via singlet oxygen-dependent processes (i.e. Type II mechanism), but rather by direct sensitizer-substrate interactions (Type I mechanisms).
  • Chem., 23:420- 429 clearly demonstrate the effects of 7,8-dimethyl-lO-ribityl isoalloxazine are due to non- singlet oxygen oxidation of guanosine residues.
  • adenosine bases appear to be sensitive to the effects of 7,8-dimethyl-10-ribityl isoalloxazine plus UV light. This is important since adenosine residues are relatively insensitive to singlet oxygen-dependent processes.
  • 7,8-dimethyl-lO-ribityl isoalloxazine appears not to produce large quantities of singlet oxygen upon exposure to UN light, but rather exerts its effects through direct interactions with substrate (e.g., nucleic acids) through electron transfer reactions with excited state sensitizer species.
  • Figure 15 depicts an embodiment of this invention in which fluid to be decontaminated is placed in a blood bag 284 equipped with an inlet port 282, through which photosensitizer in powder form 284 is added from flask 286 via pour spout 288. Quencher 300 is also added. Shaker table 280 is activated to agitate the bag 284 to dissolve photosensitizer 290 while photoradiation source 260 is activated to irradiate the fluid and photosensitizer in bag 284. Alternatively, the bag can be provided prepackaged to contain photosensitizer and quencher and the fluid is thereafter added to the bag.
  • FIGS 16 and 17 depict an embodiment of this invention in which blood bags or other photopermeable containers used in blood component collection and storage are prepackaged to contain the photosensitizer and quencher in either dry or aqueous form.
  • the additive solutions necessary for storage of blood components are added to the blood bags either separately or together with the separated blood components.
  • the additive solutions can be prepackaged in the same or connected containers in dry or aqueous form, either alone, or together with the photosensitizers necessary for viral inactivation and desired quenchers.
  • the photosensitizer, quencher and blood component additives that are prepackaged within the bags may be in a dry powder form, a pill, capsule, tablet form, liquid form, or in various combinations thereof.
  • dry solid or dry form envisions the components being in a loose powdered state or in a solid state such as a pill, capsule, tablet, or any equivalent thereof known to one skilled in the art.
  • a first blood storage bag 1 and a second blood storage bag 2 are connected together by flexible tubing 3.
  • the first and second bags 1 and 2 could also have a small container 4 located between the two blood bags via flexible tubing 3, as shown in Fig. 17.
  • the container 4 could be another bag, a flask, a reservoir, a small cylinder or any similar_container known in the art.
  • the small container 4 of Fig. 17 or the tubing 3 itself of Fig. 16 could contain certain forms of prepackaged components, in a manner similar to that of the two blood bags 1 and 2.
  • the photosensitizer, quencher, and either blood additive components or physiological saline are prepackaged in a first bag 1.
  • Glucose or another nutrient could optionally also be prepackaged in bag 1.
  • the blood additive components, quencher and photosensitizer may be in a dry solid or a liquid form. If dry form is used, a solution or preferably saline solution may be added to the bag through a port.
  • the resulting media containing blood component, photosensitizer, quencher, glucose and additive solution move via the flexible tubing 3 into a second bag 2 optionally containing prepackaged phosphate or other enhancer in either a dry solid or liquid form.
  • the second bag 2 is then disconnected from the first bag 1, mixed, and irradiated. It should be noted however, that either the first bag or the second bag could be irradiated as long as the irradiation is done after the addition of the photosensitizer.
  • the first bag 1 contains prepackaged additive solution either in solid or liquid form.
  • the resulting media including the blood component or components flows through the tubing 3 or small container 4 into the second bag 2.
  • phosphate in either a solid or liquid form is located within the tubing 3 or small container 4.
  • the phosphate dissolves upon contact into the mixture.
  • the media and dissolved phosphate mixture comes in contact with the prepackaged glucose, quencher and photosensitizer in bag 2, either in a solid or liquid form.
  • the second bag 2 is then disconnected from the first bag 1, mixed, and irradiated.
  • the first bag 1 may contain quencher, photosensitizer with or without additive solution, and also with or without glucose and the tubing 3 or small container 4 may contain phosphate.
  • the first bag 1 contains additive solution, the photosensitizer and quencher are in the tubing 3 or container 4, and phosphate and/or glucose is in the second bag 2. It is also contemplated that the photosensitizer and quencher are prepackaged in the first bag 1, and phosphate and/or glucose is in the tubing 3 or container 4.
  • the use of a frangible connection (not shown) between the first bag 1 and the container 4 is further envisioned for use with this invention. The frangible connector would be manually snapped to allow fluid or media to reach the constituent in the tubing 3 or container 4 when desired.
  • the additive solutions and other constituents can be prepackaged in either bag in aqueous or in dry solid form as well as within the small container 4.
  • additional phosphate and a nutrient such as glucose
  • the additive solution contains a phosphate and/or glucose it is contemplated that it may be unnecessary to add an additional amount of such constitutents.
  • the above are only a few examples and are not meant to be limiting. It is understood that other combinations of the constituents are also contemplated.
  • a platelet concentrate was mixed with the platelet additive solution Isolyte S at a ratio of 10:90 platelet concentrate: Isolyte S.
  • Mixtures of platelet concentrates and platelet additive solutions are referred to herein as in "media.”
  • Platelet concentrate without additive solution is referred to herein as in "plasma.”
  • Example 2 To plasma concentrate as described in Example 1 contained in a standard blood bag was added 25 ⁇ M 7,8-dimethyl-lO-ribityl isoalloxazine in powder form. The bag was spiked with bacteria as shown in Table 1, agitated and exposed to 120 J/cm 2 radiation. Inactivation results are set forth in Table 1. Table 1
  • Example 2 To platelet concentrate as described in Example 1 was added 7,8-dimethyl-lO-ribityl isoalloxazine, alloxazine mononucleotide, or 7-8-dimethyl alloxazine, followed by spiking with S. aureus or S. epidermidis, and irradiation at 80 J/cm . Inactivation results are shown in Table 2.
  • Example 2 To platelet concentrate of Example 1 was added 10 ⁇ M 7,8-dimethyl-10-ribityl- isoalloxazine. Aliquots contained no additive, 10 mM ascorbate or 10 mM KI as a
  • Example 5 To platelet concentrates of Example 1 were added varying concentrations of 7,8- dimethyl-10-ribityl-isoalloxazine. These solutions were spiked with herpes simplex virus type II (HSV-II), a double-stranded DNA envelope virus. Irradiation was done at 80 J/cm 2 . The experiment was replicated three times. In all three trials complete inactivation was achieved. Results are shown in Figure 3.
  • HSV-II herpes simplex virus type II
  • Example 5 The protocol of Example 5 was followed using S. epidermidis instead of HSV II at energies of irradiation of 40, 80 and 120 J/cm . Inactivation results are shown in Figure 4.
  • Example 7 The protocol of Example 5 was followed using ⁇ XI 74, a single stranded DNA bacteriophage, at varying concentrations of 7,8-dimethyl-lO-ribityl-isoalloxazine and energies of irradiation. Inactivation results are shown in Figure 5.
  • Example 8 To platelet concentrates of Example 1 was added 10 ⁇ M 7,8-dimethyl-lO-ribityl- isoalloxazine. These were spiked with S. aureus or ⁇ XI 74 and irradiated at varying energies of irradiation with a 50:50 mixture of visible and ultraviolet light. Inactivation results are shown in Figure 6.
  • Example 8 The protocol of Example 8 was followed using S. epidermidis and HSV-II as the microorganisms. A 50:50 mixture of ultraviolet and visible light was supplied by DYMAX light source. Inactivation results are shown in Figure 7.
  • Example 1 To platelet concentrate of Example 1 was added 10 ⁇ M 7, 8 -dimethyl- 10-ribityl- isoalloxazine in powdered form. Tests with and without added ascorbate were conducted. 150 ml of the test solutions were placed in a SpectraTM blood bag and shaken and exposed to varying energies of irradiation using 50:50 visible:ultraviolet light. After receiving 40 J/cm 2 , the contents of each bag were transferred to a new bag to avoid errors due to microorganisms which may have remained in the spike port of the bag. Inactivation results are shown in Figure 8. Downward arrows indicate inactivation to the level it was possible to detect (2.5 log titre). Example 11.
  • Example 1 To platelet concentrate of Example 1 and platelet concentrate in Isolyte S at 30:70 platelet concentrate:Isolyte S, was added 20 ⁇ M 7,8-dimethyl-10-ribityl-isoalloxazine. These were spiked with vaccinia virus, a double stranded DNA envelope virus, and exposed to 60
  • Example 13 To samples of platelet concentrate as described in Example 1 were added 5 ⁇ M or 50 ⁇ M 7,8-dimethyl-lO-ribityl-isoalloxazine. Samples were spiked with HIV 1. Using the cuvette flow cell shown in Figure 1, samples were irradiated with 50:50 visible:UV light at varying energies using an EFOS light system. Inactivation results are shown in Figure 11.
  • HIN-infected ACH-2 cells were added to samples of platelet concentrate described in Example 1. 5 or 50 ⁇ M of 7,8-dimethyl-lO-ribityl-isoalloxazine were added to the samples. The protocol of Example 13 was followed, and inactivation results are shown in Figure 12. The presence of HIV was assayed by its cytopathic effect on test cells.
  • Example 14 The protocol of Example 14 was followed and the presence of HIN assayed by quantifying the level of P24 antigen production. Inactivation results are shown in Figure 13. Example 16.
  • This example compares novel blood component additive solutions for addition to platelets separated from whole blood.
  • Six commercially available solutions were used: PAS II, PSMI-pH, PlasmaLyte A, SetoSol, PAS m, and PAS (designated PSS 1-6, respectively, in Table 4).
  • PAS II PAS II, PSMI-pH, PlasmaLyte A, SetoSol, PAS m, and PAS (designated PSS 1-6, respectively, in Table 4).
  • an effective amount of an endogenous photosensitizer 7,8-dimethyl-lO-ribityl isoalloxazine and an effective amount of a quencher.
  • the photosensitizer may be present in the various solutions at any desired concentration from about 1 ⁇ M up to the solubility of the photosensitizer in the fluid, or dry medium, and preferably about 10 ⁇ M.
  • a concentration range between about 1 ⁇ M and about 160 ⁇ M is preferred, preferably about 10 ⁇ M.
  • the composition of each solution is shown in Table 4 below, and varies in the amount of blood component additives present.
  • the blood additive components may be in a physiological solution, as well as a dry medium adapted to be mixed with a solvent, including tablet, pill or capsule form.
  • the platelet storage solution PSS 1 comprises a physiological saline solution, tri-sodium citrate at a concentration of approximately about 10 mM, sodium acetate at a concentration of approximately about 30 mM, 7, 8-dimethyl-10-ribityl isoalloxizine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • the platelet storage solution PSS 2 comprises a physiological saline solution, potassium chloride at a concentration of approximately about 5 mM, tri-sodium citrate at a concentration of approximately about 23 mM, a mixture of monosodium phosphate and dibasic sodium phosphate at a concentration of approximately about 25 mM, and 7, 8-dimethyl- 10-ribityl isoalloxizine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • the platelet storage solution PSS 3 comprises a physiological saline solution, potassium chloride at a concentration of approximately about 5 mM, magnesium chloride at a concentration of approximately about 3 mM, tri-sodium citrate at a concentration of approximately about 23 mM, sodium acetate at a concentration of approximately about 27 mM, sodium gluconate at a concentration of approximately about 23 mM, 7, 8-dimethyl- 10- ribityl isoalloxizine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • the platelet storage solution PSS 4 comprises a physiological saline solution, potassium chloride at a concentration of approximately about 5 mM, magnesium chloride at a concentration of approximately about 3 mM, tri-sodium citrate at a concentration of approximately about 17 mM, sodium phosphate at a concentration of approximately about 25 mM, sodium acetate at a concentration of approximately about 23 mM, glucose at a concentration of approximately about 23.5 mM, maltose at a concentration of approximately about 28.8 mM, 7, 8-dimethyl- 10-ribityl isoalloxizine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • potassium chloride at a concentration of approximately about 5 mM
  • magnesium chloride at a concentration of approximately about 3 mM
  • tri-sodium citrate at a concentration of approximately about 17 mM
  • sodium phosphate at a concentration of approximately about 25 mM
  • sodium acetate at a concentration of approximately about 23
  • the platelet storage solution PSS 5 comprises a physiological saline solution, potassium chloride at a concentration of approximately about 5.1 mM, calcium chloride at a concentration of approximately about 1.7 mM, magnesium sulfate at a concentration of approximately about 0.8 mM, tri-sodium citrate at a concentration of approximately about 15.2 mM, citric acid at a concentration of approximately about 2.7 mM, sodium bicarbonate at a concentration of approximately about 35 mM, sodium phosphate at a concentration of approximately about 2.1 mM, glucose at a concentration of approximately about 38.5 mM, 7,8-dimethyl- 10-ribityl isoalloxizine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • potassium chloride at a concentration of approximately about 5.1 mM
  • calcium chloride at a concentration of approximately about 1.7 mM
  • magnesium sulfate at a concentration of approximately about 0.8 mM
  • tri-sodium citrate
  • the platelet storage solution PSS 6 comprises a physiological saline solution, tri-sodium citrate at a concentration of approximately about 12.3 mM, sodium phosphate at a concentration of approximately about 28 mM, sodium acetate at a concentration of approximately about 42 mM, 7,8-dimethyl-lO-ribityl isoalloxizine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • physiologic saline may be replaced with a solvent comprising water and an effective amount of sodium chloride.
  • the blood additive solution would comprise a commercially available product for example PAS II or T-Sol (which has the same ingredients as PAS II) and an effective amount of a nutrient such as glucose, an enhancer such as phosphate, 7,8-dimethyl-lO-ribityl isoalloxazine in a pill or a dry medium form, and an effective amount of a quencher.
  • a nutrient such as glucose
  • an enhancer such as phosphate, 7,8-dimethyl-lO-ribityl isoalloxazine in a pill or a dry medium form
  • a quencher an effective amount of a quencher.
  • the blood additive solution could comprise an other additive solution including an effective amount of 7, 8-dimethyl- 10-ribityl isoalloxazine in a liquid, pill or dry medium form.
  • PSS 7, PSS 8 and PSS 9 are examples of such blood additive solutions set forth in Table 5 below.
  • PSS 7 was prepared in RODI water and sodium chloride at a concentration of approximately 115 mM, sodium citrate at a concentration of approximately 10.0 mM, sodium phosphate (monobasic) at a concentration of approximately 6.2 mM, sodium phosphate (dibasic) at a concentration of approximately 19.8 mM, sodium acetate at a concentration of approximately 30.0 mM, 7,8-dimethyl 10-ribityl isoalloxazine at a concentration of approximately 14.0 ⁇ M, and an effective amount of a quencher.
  • the solution has a pH of 7.2.
  • PSS 8 was prepared in RODI water and comprises and sodium chloride at a concentration of approximately 78.3 mM, potassium chloride at a concentration of approximately 5.7 mM, magnesium chloride at a concentration of approximately 1.7 mM, sodium phosphate (monobasic) at a concentration of approximately 5.4 mM, sodium phosphate (dibasic) at a concentration of approximately 24.6 mM, sodium acetate at a concentration of approximately 34.3 mM, a variable concentration of 7,8-dimethyl 10-ribityl isoalloxazine, and an effective amount of a quencher.
  • the solution has a pH of 7.4, and an osmolarity of 297 mmol/kg.
  • PSS 9 was prepared in RODI water and comprises and sodium chloride at a concentration of approximately 68.5 mM, potassium chloride at a concentration of approximately 5.0 mM, magnesium chloride at a concentration of approximately 1.5 mM, sodium phosphate (monobasic) at a concentration of approximately 8.5 mM, sodium phosphate (dibasic) at a concentration of approximately 21.5 mM, sodium acetate at a concentration of approximately 30.0 mM, 7,8-dimethyl 10-ribityl isoalloxazine at a concentration of approximately 14.0 ⁇ M, and an effective amount of a quencher.
  • the solution has a pH of 7.2, and an osmolarity of 305 mmol/kg.
  • This example compares blood component additive solutions for addition to red blood cells separated from whole blood.
  • AS-1, AS-3, AS-5, SAGM, and MAP designated AS1, AS2, AS3, AS4 and AS5, respectively in Table 6
  • MAP designated AS1, AS2, AS3, AS4 and AS5, respectively in Table 6
  • an effective amount of an endogenous photosensitizer 7,8-dimethyl-lO-ribityl isoalloxazine
  • an effective amount of a quencher The photosensitizer may be present in the various solutions at any desired concentration from about 1 ⁇ M up to the solubility of the photosensitizer in the fluid, or dry medium, and preferably about 10 ⁇ M.
  • the quencher may be present in a range of 0.05 mM up to a concentration of about lOOmM. The preferred range is between about 0.1 to about 20 mM. For 7,8-dimethyl-lO-ribityl isoalloxazine a concentration range between about 1 ⁇ M and about 160 ⁇ M is preferred, preferably about 10 ⁇ M.
  • the composition of each additive solution is shown in Table 6 below, and varies in the amount of blood component additives present.
  • the red blood cell additive solution components may be in a physiological solution, a dry medium adapted to be mixed with a solvent, including in tablet, pill or capsule form as described above.
  • the red blood cell additive solution AS 1 comprises a physiological saline solution, dextrose at a concentration of approximately about 122.09 mM, adenine at a concentration of approximately about 2 mM, mannitol at a concentration of approximately about 41.16 mM, 7,8-dimethyl- 10-ribityl isoalloxazine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • the red blood cell additive solution AS 2 comprises a physiological saline solution, dextrose at a concentration of approximately about 61.04 mM, adenine at a concentration of approximately about 2.22 mM, sodium phosphate (monobasic) at a concentration of approximately about 23 mM, sodium citrate at a concentration of approximately 19.99, and citric acid at a concentration of about 2.19 mM, 7,8-dimethyl-lO- ribityl isoalloxazine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • dextrose at a concentration of approximately about 61.04 mM
  • adenine at a concentration of approximately about 2.22 mM
  • sodium phosphate (monobasic) at a concentration of approximately about 23 mM
  • sodium citrate at a concentration of approximately 19.99
  • citric acid at a concentration of about 2.19 mM
  • the red blood cell additive solution AS 3 comprises a physiological saline solution, dextrose at a concentration of approximately about 49.94 mM, adenine at a concentration of approximately about 2.22 M, mannitol at a concentration of approximately about 28.81 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • the red blood cell additive solution AS 4 comprises a physiological saline solution, dextrose at a concentration of approximately about 49.94 mM, adenine at a concentration of approximately about 1.25 mM, mannitol at a concentration of approximately about 28.81 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • the red blood cell additive solution AS 5 comprises a physiological saline solution, adenine at a concentration of approximately about 0.10 mM, sodium phosphate (monobasic) at a concentration of approximately about 7.80 mM, mannitol at a concentration of approximately about 79.91 mM, sodium citrate at a concentration of approximately about 6 mM, glucose at a concentration of approximately about 39.96 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • a physiological saline solution adenine at a concentration of approximately about 0.10 mM
  • sodium phosphate (monobasic) at a concentration of approximately about 7.80 mM
  • mannitol at a concentration of approximately about 79.91 mM
  • sodium citrate at a concentration of approximately about 6 mM
  • glucose at a concentration of approximately about 39.96 mM
  • the present invention further contemplates the addition of a red blood cell anticoagulant-based solution to the separated red blood cells.
  • a red blood cell anticoagulant-based solution Five commercially available anticoagulant-based solutions were considered: CPD, CP2D, CPDA-1, ACD-A, and ACD-B (designated ABS1, ABS2, ABS3, ABS4 and ABS5 respectively, in Table 7).
  • CPD CPD
  • CP2D CPD
  • CPDA-1 CPDA-1
  • ACD-A CPDA-1
  • ACD-A designated ABS1, ABS2, ABS3, ABS4 and ABS5 respectively, in Table 7
  • an effective amount of an endogenous photosensitizer 7,8-dimethyl-lO- ribityl isoalloxazine and an effective amount of a quencher.
  • the photosensitizer may be present in the various solutions at any desired concentration from about 1 ⁇ M up to the solubility of the photosensitizer in the fluid, or dry medium, and preferably about 10 ⁇
  • a concentration range between about 1 ⁇ M and about 160 ⁇ M is preferred, preferably about 10 ⁇ M.
  • the composition of each solution is shown in Table 7 below, and varies in the amount of blood component additives present.
  • the blood additive components may be in a physiological solution, a dry medium adapted to be mixed with a solvent, or in tablet, pill or capsule form as described above.
  • the red blood cell anticoagulant-based solution ABS 1 comprises a physiological saline solution, sodium citrate at a concentration of approximately about 89.59 mM, citric acid at a concentration of approximately about 15.53 mM, dextrose at a concentration of approximately about 141.82 mM, sodium phosphate monobasic at a concentration of approximately about 18.52 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • the red blood cell anticoagulant-based solution ABS 2 comprises a physiological saline solution, sodium citrate at a concentration of approximately about 89.59 mM, citric acid at a . concentration of approximately about 15.53 mM, dextrose at a concentration of approximately about 283.64 mM, sodium phosphate monobasic at a concentration of approximately about 18.52 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • the red blood cell anticoagulant-based solution ABS 3 comprises a physiological saline solution, sodium citrate at a concentration of approximately about 89.59 mM, citric acid at a concentration of approximately about 15.53 mM, dextrose at a concentration of approximately about 177.05 mM, sodium phosphate monobasic at a concentration of approximately about 18.52 mM, adenine at a concentration of approximately about 2.03 mM, 7,8-dimethyl- 10-ribityl isoalloxazine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • the red blood cell anticoagulant-based solution ABS 4 comprises a physiological saline solution, dextrose at a concentration of approximately about 135.96 mM, sodium citrate at a concentration of approximately about 74.80 mM, citric acid at a concentration of approximately about 41.64 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • the red blood cell anticoagulant-based solution ABS 5 comprises a physiological saline solution, dextrose at a concentration of approximately about 81.58 mM, sodium citrate at a concentration of approximately about 44.88 mM, citric acid at a concentration of approximately about 24.99 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M, and an effective amount of a quencher.
  • This example compares blood component additive solutions for addition to red blood cells separated from whole blood.
  • Five commercially available red blood cell additive solutions were considered: AS-1, AS-3, AS-5, SAGM, and MAP. These solutions are listed as (1-5) in Table 8.
  • an effective amount of an endogenous photosensitizer, 7,8-dimethyl-lO-ribityl isoalloxazine and an effective amount of a quencher was added to each known solution was added an effective amount of an endogenous photosensitizer, 7,8-dimethyl-lO-ribityl isoalloxazine and an effective amount of a quencher.
  • the photosensitizer may be present in the various solutions at any desired concentration from about 1 ⁇ M up to the solubility of the photosensitizer in the fluid, or dry medium, and preferably about 10 ⁇ M.
  • a concentration range between about 1 ⁇ M and about 160 ⁇ M is preferred, preferably about 10 ⁇ M.
  • the composition of each additive solution is shown in Table 8 below, and varies in the amount of blood component additives present.
  • the red blood cell additive solution components may be in a physiological solution, a dry medium adapted to be mixed with a solvent, including in tablet, pill or capsule form as described above.
  • the red blood cell additive solution 1 (Commercial Solution AS-1) comprises a physiological saline solution, dextrose at a concentration of approximately about 122.09 mM, adenine at a concentration of approximately about 2 mM, mannitol at a concentration of approximately about 41.16 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M and a variable amount of at least one quencher.
  • Communication Solution AS-1 comprises a physiological saline solution, dextrose at a concentration of approximately about 122.09 mM, adenine at a concentration of approximately about 2 mM, mannitol at a concentration of approximately about 41.16 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M and a variable amount of at least one quencher.
  • the red blood cell additive solution 2 (Commercial Solution AS-3) comprises a physiological saline solution, dextrose at a concentration of approximately about
  • adenine at a concentration of approximately about 2.22 mM
  • sodium phosphate monobasic
  • sodium citrate at a concentration of approximately 19.99
  • citric acid at a concentration of about 2.19 mM
  • 7,8- dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M and a variable concentration of at least one quencher.
  • the red blood cell additive solution 3 (Commercial Solution AS-5) comprises a physiological saline solution, dextrose at a concentration of approximately about 49.94 mM, adenine at a concentration of approximately about 2.22 mM, mannitol at a concentration of approximately about 28.81 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M and a variable concentration of at least one quencher.
  • the red blood cell additive solution 4 (Commercial Solution SAGM) comprises a physiological saline solution, dextrose at a concentration of approximately about 49.94 mM, adenine at a concentration of approximately about 1.25 mM, mannitol at a concentration of approximately about 28.81 mM, 7,8-dimethyl-10- ⁇ bityl isoalloxazine at a concentration of about 10 ⁇ M and a variable concentration of at least one quencher.
  • Communication Solution SAGM comprises a physiological saline solution, dextrose at a concentration of approximately about 49.94 mM, adenine at a concentration of approximately about 1.25 mM, mannitol at a concentration of approximately about 28.81 mM, 7,8-dimethyl-10- ⁇ bityl isoalloxazine at a concentration of about 10 ⁇ M and a variable concentration of at least one quencher.
  • the red blood cell additive solution 5 (Commercial Solution MAP) comprises a physiological saline solution, adenine at a concentration of approximately about 0.10 mM, sodium phosphate (monobasic) at a concentration of approximately about 7.80 mM, mannitol at a concentration of approximately about 79.91 mM, sodium citrate at a concentration of approximately about 6 mM, glucose at a concentration of approximately about 39.96 mM, 7,8-dimethyl-lO-ribityl isoalloxazine at a concentration of about 10 ⁇ M and a variable concentration of at least one quencher.
  • adenine at a concentration of approximately about 0.10 mM
  • sodium phosphate monobasic
  • mannitol at a concentration of approximately about 79.91 mM
  • sodium citrate at a concentration of approximately about 6 mM
  • glucose at a concentration of approximately about 39.96 mM
  • the photosensitizer and quencher may also be added to the cells to be pathogen inactivated in a sterile saline or buffer solution without other additives.
  • each unit of red blood cells was separated from its plasma and diluted to a hematocrit of 30%.
  • Table 9 shows results of the DAT tests (as either + or -). Table 9 also compares the percentage of hemolysis of whole blood, red blood cells only after plasma expression and dilution with riboflavin and quencher (designated post-dilution), and after 50 minutes of illumination.
  • Figure 18 is a graph of BVDV kill in red blood cells using different treatment conditions over time.
  • GSH stands for reduced glutathione
  • N-AC stands for N-acetyl cysteine.
  • Red blood cells for the conditions “+GSH”, “+GSH + mannitol” and “+N-AC” were prepared by plasma expression, followed by dilution with 500 uM riboflavin and 4 mM quencher.
  • Red cells for the condition labelled "1-wash” were prepared by plasma expression, resuspension in normal saline, recentrifugation and removal of wash solution, then dilution with 500 uM riboflavin and 4 mM GSH.
  • 2991 wash were washed with the 2991 wash the equivalent of 3 manual washes, then diluted in 500 ⁇ M riboflavin for illumination (no quencher was added).
  • a 2991 wash refers to washing the red blood cells using a 2991 cell processing machine available from Gambro BCT, Inc. (Lakewood, CO, USA.). The 2991 washes the red cells with 700 mL of 0.9% NaCl and 300 mL of 500 ⁇ M riboflavin in 0.9% NaCl.
  • the product of the wash step is a suspension of concentrated RBCs at a 60 to 70% hematocrit with a riboflavin concentration of approximately 370 ⁇ M.
  • a calculated volume of the washed red cells is mixed with 550 ⁇ M riboflavin in 0.9% NaCl in the ELP bag to obtain a suspension with a hematocrit of 50% and a volume of 276 mL.
  • the 2991 wash removed extracellular proteins from the red cell suspension. When red cells are washed thoroughly prior to illumination, DAT results for red cells after illumination are negative. An additional benefit of the wash to the red cell process is increased rate of vi s kill.
  • the wash step also provides some virus removal.
  • a 4 log BVDV viral kill may be obtained at 25-30 minutes of light exposure by expressing plasma from the collected red blood cells before the addition of the irradiation solution containing 4 mM glutathione and 500 M riboflavin.
  • Figure 19 is a graph comparing the log reduction of BVDV in washed red blood cells as compared to non-washed red blood cells. As can be seen from the figure, there is a greater variability in results obtained from the unwashed red blood cells due to the presence of plasma proteins in the unwashed red blood cells.
  • the abbreviations and other conditions used in Figure 19 are the same as in Figure 18. Mannitol was also added.
  • Figure 20 is a graph indicating the inactivation kinetics of BVDV virus in the presence and absence of Vitamin E in platelets.
  • Figure 20 shows log virus titre. 419 nm light was used with 30 uM riboflavin, with 120 cpm linear mixing.
  • Figure 21 is a graph indicating the level of inactivation observed between platelet samples with and without Vitamin E.
  • Figure 22 is a graph showing the results of survival of radiolabelled primate platelets under varying treatment conditions (with and without antioxidants and untreated controls). The results indicate that there is a substantial improvement (back to untreated control levels) in the recovery of cells observed with the addition of vitamin E and vitamin C.
  • Table 10 shows DAT results of red blood cells incubated with a quencher and riboflavin and illuminated at 190 J/cm 2 .
  • the cells were not washed prior to the addition of quencher and riboflavin.
  • the cells were illuminated in a 150 mL ELP bag in a volume of 77 mLs solution and 38 mLs of air.
  • the red blood cells were diluted to a hematocrit of 30%.
  • the quenchers either N-acetyl-L-cysteine (NAC) or glutathione (GSH) were added to each unit in varying concentrations in 500 uM riboflavin and 0.9% saline.

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Abstract

L'invention concerne des méthodes et des appareils destinés à l'inactivation de micro-organismes dans des fluides ou sur des surfaces. De préférence, ces fluides contiennent du sang ou des produits sanguins ainsi que des protéines biologiquement actives. Des méthodes préférées d'inactivation de micro-organismes consistent à ajouter une dose efficace non toxique d'un photosensibilisant endogène et d'un extincteur dans un fluide, puis à exposer ce fluide à un photorayonnement suffisamment puissant pour activer le photosensibilisant endogène, d'où l'inactivation des micro-organismes. L'extincteur réduit les réactions secondaires produites par le photosensibilisant, la lumière pouvant alors endommager les constituants biologiques souhaités.
EP02744213A 2001-05-30 2002-05-30 Procede d'inactivation virale utilisant un antioxydant Withdrawn EP1404379A2 (fr)

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TW590780B (en) * 2000-06-02 2004-06-11 Gambro Inc Additive solutions containing riboflavin
JP2007532606A (ja) 2004-04-16 2007-11-15 ヘルボ・フオトデユナーミツク・システムズ・ゲー・エム・ベー・ハー・ウント・コー・カー・ゲー 微生物の光力学的抑制のための調製物及び該調製物の使用
ES2401148T3 (es) 2004-04-16 2013-04-17 Bredent Medical Gmbh & Co. Kg Preparación para combatir fotodinámicamente microorganismos y uso de la preparación
CN101360747B (zh) * 2006-01-27 2011-08-10 科安比司特生物技术有限责任公司 生产高浓度咯嗪溶液的方法和组合物
US8580192B2 (en) 2006-10-31 2013-11-12 Ethicon, Inc. Sterilization of polymeric materials
WO2009018309A2 (fr) * 2007-08-01 2009-02-05 Caridianbct Biotechnologies, Llc Inactivation de pathogènes du sang entier
US12089589B2 (en) 2009-10-12 2024-09-17 Hemanext Inc. Irradiation of red blood cells and anaerobic storage
US9199016B2 (en) 2009-10-12 2015-12-01 New Health Sciences, Inc. System for extended storage of red blood cells and methods of use
US11284616B2 (en) 2010-05-05 2022-03-29 Hemanext Inc. Irradiation of red blood cells and anaerobic storage
EP2729000B1 (fr) * 2011-07-05 2022-12-14 Hemanext Inc. Un procédé pour le stockage prolongé des érythrocytes
EA034347B1 (ru) * 2011-10-26 2020-01-30 Амген Инк. Способ инактивации вирусов при получении антител
KR102701691B1 (ko) 2015-03-10 2024-08-30 헤마넥스트 인코포레이티드 산소 감소 1회용 키트, 장치 및 이의 사용 방법
WO2018201065A1 (fr) * 2017-04-27 2018-11-01 Fred Hutchinson Cancer Research Center Formules thérapeutiques contenant des cellules souches cd34 + dérivées de la sélection négative

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US6258577B1 (en) * 1998-07-21 2001-07-10 Gambro, Inc. Method and apparatus for inactivation of biological contaminants using endogenous alloxazine or isoalloxazine photosensitizers
US6268120B1 (en) * 1999-10-19 2001-07-31 Gambro, Inc. Isoalloxazine derivatives to neutralize biological contaminants

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