[go: up one dir, main page]

WO2007027178A2 - Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts or repair - Google Patents

Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts or repair Download PDF

Info

Publication number
WO2007027178A2
WO2007027178A2 PCT/US2005/031117 US2005031117W WO2007027178A2 WO 2007027178 A2 WO2007027178 A2 WO 2007027178A2 US 2005031117 W US2005031117 W US 2005031117W WO 2007027178 A2 WO2007027178 A2 WO 2007027178A2
Authority
WO
WIPO (PCT)
Prior art keywords
platelets
wound healant
wound
plasma
thrombin
Prior art date
Application number
PCT/US2005/031117
Other languages
French (fr)
Other versions
WO2007027178A3 (en
Inventor
Mackie J. Walker, Jr.
James Bennie Gandy
Original Assignee
Walker Mackie J Jr
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 Walker Mackie J Jr filed Critical Walker Mackie J Jr
Priority to PCT/US2005/031117 priority Critical patent/WO2007027178A2/en
Priority to CA002621140A priority patent/CA2621140A1/en
Priority to EP05793313A priority patent/EP1943349A2/en
Priority to BRPI0520502-6A priority patent/BRPI0520502A2/en
Publication of WO2007027178A2 publication Critical patent/WO2007027178A2/en
Publication of WO2007027178A3 publication Critical patent/WO2007027178A3/en

Links

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
    • A61K31/56—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids
    • 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
    • A61K31/045—Hydroxy compounds, e.g. alcohols; Salts thereof, e.g. alcoholates
    • A61K31/07—Retinol compounds, e.g. vitamin A
    • 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
    • A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
    • A61K31/19—Carboxylic acids, e.g. valproic acid
    • A61K31/195—Carboxylic acids, e.g. valproic acid having an amino group
    • A61K31/197—Carboxylic acids, e.g. valproic acid having an amino group the amino and the carboxyl groups being attached to the same acyclic carbon chain, e.g. gamma-aminobutyric acid [GABA], beta-alanine, epsilon-aminocaproic acid or pantothenic acid
    • 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
    • A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
    • A61K31/19—Carboxylic acids, e.g. valproic acid
    • A61K31/20—Carboxylic acids, e.g. valproic acid having a carboxyl group bound to a chain of seven or more carbon atoms, e.g. stearic, palmitic, arachidic acids
    • A61K31/203—Retinoic acids ; Salts thereof
    • 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
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
    • A61K31/352—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom condensed with carbocyclic rings, e.g. methantheline 
    • A61K31/353—3,4-Dihydrobenzopyrans, e.g. chroman, catechin
    • A61K31/355—Tocopherols, e.g. vitamin E
    • 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
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/365—Lactones
    • A61K31/375—Ascorbic acid, i.e. vitamin C; Salts thereof
    • 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
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
    • A61K31/403—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil condensed with carbocyclic rings, e.g. carbazole
    • A61K31/404—Indoles, e.g. pindolol
    • A61K31/4045—Indole-alkylamines; Amides thereof, e.g. serotonin, melatonin
    • 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
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
    • A61K31/7034—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin
    • A61K31/7036—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin having at least one amino group directly attached to the carbocyclic ring, e.g. streptomycin, gentamycin, amikacin, validamycin, fortimicins
    • 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
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7042—Compounds having saccharide radicals and heterocyclic rings
    • A61K31/7052—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides
    • A61K31/706—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom
    • A61K31/7064—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom containing condensed or non-condensed pyrimidines
    • A61K31/7076—Compounds having saccharide radicals and heterocyclic rings having nitrogen as a ring hetero atom, e.g. nucleosides, nucleotides containing six-membered rings with nitrogen as a ring hetero atom containing condensed or non-condensed pyrimidines containing purines, e.g. adenosine, adenylic acid
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00—Medicinal preparations containing inorganic active ingredients
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K33/00—Medicinal preparations containing inorganic active ingredients
    • A61K33/06—Aluminium, calcium or magnesium; Compounds thereof, e.g. clay
    • 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
    • A61K35/16—Blood plasma; Blood serum
    • 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
    • A61K35/19—Platelets; Megacaryocytes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/39—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin, cold insoluble globulin [CIG]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/43—Enzymes; Proenzymes; Derivatives thereof
    • A61K38/46—Hydrolases (3)
    • A61K38/48—Hydrolases (3) acting on peptide bonds (3.4)
    • A61K38/482—Serine endopeptidases (3.4.21)
    • A61K38/4833—Thrombin (3.4.21.5)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca

Definitions

  • the invention relates to an improved Lyophilized platelet rich plasma used to make a platelet gel wound healant, and methods of preparation and use thereof for healing wounds are disclosed.
  • the improved wound healant comprises a therapeutically effective amount of activated growth factors with optional none, one or more additional anti-oxidant such as vitamin A and/or C and/or E, and/or one or more antibiotics and/or GHK-Cu (produced by
  • the present invention relates to improved Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts and methods of making and use thereof of Lyophilized or fixed platelets.
  • the present invention generally relates to the therapeutic uses of blood platelets and fresh plasma that has been lyophilized, and more particularly to manipulations or modifications of platelets and plasma, such as in preparing freeze-dried compositions that can be rehydrated at the time of application and which when rehydrated have a normal response to thrombin and other agonists with respect to that of fresh platelets.
  • the inventive compositions are useful in applications such as wound care.
  • the platelets may be suspended, for example, in a solution containing a cryoprotectant at a temperature of about 22.degree. C. and then cooled to below 15. degree. C. This incorporates some cryoprotectant into the cells.
  • a wound is generally defined as an injury to an area of the body of a human or animal. Although injury to the surface of the skin is the most well known type of wound, the surfaces of internal organs may also be wounded, such as during surgery, rupture of the spleen or liver, or resulting from traumatic blows to the body surface in the vicinity of an internal organ.
  • Tissue wounds may have a wide spectrum of manifestations, as small as merely an abnormal microscopic tear or fissure in tissue (or a surface thereof), or as large as the abrasion or ablation of the skin covering a substantial portion of the body, such as in a burn victim.
  • Acute wounds covering a large or movable surface are usually the most difficult to guard from infection, and to heal.
  • Blood and bodily fluids include various substances that affect wound healing.
  • the blood is the primary medium for delivering healing agents to the wound site, and for transporting foreign or harmful substances away from the wound.
  • Whole blood is primarily comprised of three main types of cells suspended in a protein rich solution known as plasma.
  • the three main cell types in whole blood are erythrocytes (a.k.a. red blood cells), leukocytes (a.k.a. white blood cells) and thrombocytes (a.k.a. platelets).
  • the red blood cells are the iron-containing cells that facilitate the transport and transfer of oxygen to body tissue, and the removal of carbon dioxide.
  • the white blood cells perform a variety of functions such as phagocytosis of foreign bodies and production of antibodies, and are primarily responsible for fighting infection and foreign substances within the blood or wound site. Platelets perform many functions such as plugging leaks in blood vessels and helping begin the process leading to the formation of a blood clot; platelets contain substances known as growth factors that facilitate the formation of new tissue.
  • apheresis apheresis
  • the red and white blood cells and plasma may be separated out and returned to the donor's or patient's body, leaving the sequestered platelets in essentially concentrated form for use in wound healing techniques. This may be preformed at a blood collection center, blood bank, at the Physicians Clinic or Hospital. From blood extracted from a patient or from the other forms of collection methods, the platelets may thus be obtained and activated for use on the patient; methods of using a patient's own blood are called “autologous" or "autogenic" donor methods.
  • Another method using blood donated by one or more third parties for use by a patient are called “homologous” or “heterologous” donor methods, or collectively called “allogenic” methods.
  • One of the proteins suspended in plasma is fibrinogen, which reacts with substances released into (or attracted by) wound sites to produce sticky strands of fibrin. Such reactions result in the cross linking of the fibrin strands to form a mesh that holds and supports the deposit or growth of other tissue materials at the wound site. Therefore, the need for fresh plasma also known in the art "The Plasma Back Bone”.
  • the wound healing process is generally considered to occur in several stages, generally known as the healing cascade.
  • platelets are among the first cells to appear in the vicinity of the wound.
  • Activation of a platelet by an agonist such as thrombin, or other agonists such as those listed elsewhere herein leads to the release of granule material from within the platelet.
  • Such granulation activation results in the release of proteins known as growth factors, primarily concentrated in the alpha granules of platelets.
  • growth factors primarily concentrated in the alpha granules of platelets.
  • These released growth factors stimulate the formation of new tissue; when applied to wounds, growth factors have been known to increase the rate of collagen laydown, vascular ingrowth, fibroblast proliferation and overall healing.
  • PDGF platelet-derived growth factor
  • fibrin glue which often are essentially a mixture of co-coagulants (thrombin and calcium), concentrated fibrinogen and other coagulation proteins.
  • co-coagulants thrombin and calcium
  • fibrin glue the primary roles of fibrin glue are to seal wound surfaces to prevent loss of blood and other body fluids after surgery, and to provide adhesion between adjacent tissue surfaces. These products form a hard, cast-like covering over the area to be sealed, and tend to be non- yielding to limb movement. While there has been much research concerning fibrin glue, this material belongs to a separate field from the present invention, primarily because fibrin glues typically contain cryoprecipitated proteins without platelets. The use of fibrin glue is discussed extensively in the scientific literature; for example, see the references cited in U.S. Pat. No.
  • Wound treatment compositions derived from platelet enriched concentrates are known and possess certain advantages over materials without platelets such as fibrin glue.
  • One reason is that natural wound healing agents are released by the platelets.
  • the concentration of platelets likewise allows for a concentrated amount of wound healing factors.
  • Representative examples of platelet derived wound treatment compositions are described for instance in Hood U.S. Pat. No. 5,733,545; Knighton U.S. Pat. No. 5,165,938; and Gordiner U.S. Pat. No. 5,599,558.
  • This form of treatment has major problems, like time consuming and problems collecting blood sometimes with expensive machines and disposables.
  • Platelet concentrates are typically isolated by the process of differential centrifugation, which essentially allows separating the blood into at least three different components: packed erythrocytes (red blood cells), plasma and platelet concentrate.
  • Platelet concentrate can be combined with a solution of either sodium or calcium mixed with thrombin ("calcified thrombin"), which instantaneously form a composition of activated platelets that, when made with the necessary viscosity, can be utilized as a wound sealant.
  • thrombin thrombin
  • the chemical reactions and cascades that normally occur when thrombin is added to the concentrated platelets are indeed complex. See, for instance, Reeder, et al, in Proceedings of the American Academy of Cardiovascular Perfusion, Vol. 14, January 1993.
  • Such wound sealants typically set up into a hard mass covering the application site, thereby sealing the site against further blood loss and external contaminants.
  • an improved Lyophilized platelet enriched plasma wound treatment composition which avoids or diminishes the problems associated with typical platelet enriched wound compositions and would be desirable.
  • a dehydrated composition comprising freeze-dried platelets and fresh plasma that are effectively to preserve biological properties during freeze-drying and rehydration.
  • These platelets are rehydratable so as to have a normal response to at least one agonist, such as thrombin.
  • substantially all freeze-dried platelets of the invention when rehydrated and mixed with thrombin (1 U/ml) form a clot within three minutes at 37.degree. C.
  • the dehydrated composition can include one or more other agents, such as antibiotics, antifungals, growth factors, or the like, depending upon the desired therapeutic application.
  • the present invention relates to an improved Lyophilized platelet and plasma composition for wound treatment, a method of making and use thereof.
  • the composition comprises Lyophilized platelets and plasma composition in liquid form to the wound site and will gel to prevent the material from flowing away from the site.
  • Optional antibiotics may be included in the improved composition to prevent infections at the wound site.
  • Lyophilized platelet gels containing fresh plasma is prepared for topical application at the wound site and avoid the requirement for daily reapplication.
  • One such method is a process of preparing a dehydrated composition comprising providing a source of platelets and fresh plasma and lyophilizing the platelets and plasma. The rehydration preferably done with de- ionized water.
  • compositions of matter and the methods described herein are not limited solely to topical application.
  • the inventive composition increases the amount of growth factors in the wound, and thereby facilitates the promotion of the healing rate. This may be especially important in "wounded" patients, especially those with chronic wounds who may lack sufficient circulation to facilitate the healing cascade.
  • the invention described herein also facilitates the covering of the wound area with a substance that prevents or helps to reduce infection caused by most bacteria.
  • Practice of the invention permits the manipulation or modification of platelets and plasma while maintaining, or preserving, biological properties, such as a response to thrombin.
  • the inventive freeze-dried platelets and plasma including the freeze-dried platelets are substantially shelf stable at ambient temperatures when packaged in moisture barrier materials.
  • the invention described herein expands the uses for concentrated platelet materials, especially those in gel form, by improving the speed and convenience of making the composition; the invention described herein also improves the performance of the concentrated platelet composition, by making it more useable for applications over longer periods of time, and by enhancing the wound healing and infection fighting properties.
  • Another aspect of the present invention involves adding one or more antibiotic substance at one or more times during the processing period so that the resulting concentrated platelet and plasma composition contains either one or a variety of the antibiotics.
  • the use of an antibiotic in concentrated platelet and plasma compositions that enhances the complex healing cascade is indeed novel.
  • the invention disclosed herein involves adding such substances in a manner that does not detract from, substantially interfere with, or even destroy these different reactions, pH balances and potency.
  • Vitamins are known to have wound healing and anti-oxidant properties.
  • vitamins include vitamin E, vitamin A, vitamin C and other retinoids.
  • non-vitamin anti-oxidants may be included in the concentrated platelet gel.
  • Non-limiting representative examples of such anti-oxidants include beta-carotene.
  • the phrase blood collecting or blood extraction includes techniques, materials and apparatus known in the field, such as (for example) inclusion of anticoagulation materials, the use of blood drawing and infusion apparatus.
  • the phrase growth factor means any material(s) promoting growth of a tissue.
  • thrombin may include calcified thrombin, in particular, about 5,000 units of thrombin per 5 ml of 10% of aqueous calcium chloride solution; it may include calcified bovine thrombin as well as autologous thrombin, allogeneic thrombin or recombinant human thrombin.
  • viscosity means those characteristics of the specified material(s) determining the degree of gelation, such as (for example) the firmness or hardness of the material, or the degree to which the material resists flowing like a fluid.
  • therapeutically effective amount means the amount or amounts of the constituent elements or combination thereof necessary to enhance wound healing such as, for example, the reduction in the volume or surface area of a wound, the increase in the amount of granulation tissue or other biological material facilitating collagen laydown, vascular ingrowth, fibroblast proliferation or overall healing; all of the versions of the invention described herein are assumed to have the therapeutically effect amount(s) of constituent substances, or combinations thereof.
  • anti-oxidant refers to any material(s) having anti-oxidant properties.
  • Anti-oxidant would include, without limitation, vitamins such as vitamins C, A and E and non-vitamins such as -carotene.
  • the invention includes a wound healant composition comprising activated growth factors and ascorbic acid.
  • said growth factors are included within platelets.
  • the body produces many substances generally known as growth factors, and these growth factors are contemplated for use in the present invention.
  • the preferred growth factors for use in the present invention are selected from the group consisting of platelet-derived growth factor (PDGF), platelet-derived angiogenesis factor (PDAF), vascular endotheial growth factor (VEGF), platelet-derived epidermal growth factor (PDEGF), platelet factor 4 (PF-4), transforming growth factor .beta.
  • TGF-B acidic fibroblast growth factor
  • FGF-A basic fibroblast growth factor
  • FGF-B transforming growth factor .alpha.
  • TGF-A insulin-like growth factors 1 and 2
  • BCG thromboglobulin-related proteins
  • TSP thrombospondin
  • vWF von Wallinbrand's factor
  • fibropeptide A fibrinogen, albumin, plasminogen activator inhibitor 1 (PAI-1 ), osteonectin, regulated upon activation normal T cell expressed and presumably secreted (RANTES), gro-.alpha., vitronectin, fibrin D-dimer, factor V, antithrombin III, immunoglobulin-G (IgG), immunoglobulin-M (IgM), immunoglobulin-A (IgA), a2-macroglobulin, angiogenin, Fg-D, elastase, keratinocyte growth
  • each substance is known or believed to enhance cell or tissue growth.
  • said substances, or various combinations thereof are known or believed to function together in an unexpected synergistic manner to promote wound healing.
  • Suitable, non- limiting, anti-oxidants useful in the invention include but are not limited to vitamins such as vitamin C (ascorbic acid), vitamin E, vitamin A and other retinoids; and the carotenes such as beta-carotene. In practicing this invention, ascorbic acid as anti-oxidant is particularly preferred.
  • the platelets are separated from the red blood cells and white blood cells of whole blood, primarily through differential centrifugation, although any suitable method for separating platelets from whole blood may be employed in practicing this invention.
  • the overall composition of the invention disclosed herein may contain incidental amounts of white blood cells, due to the fact that the platelets are rarely totally isolated from the other blood components. It is believed that the present invention contains only minimal or trace amounts of white blood cells; it is believed that the white blood cell count of the present invention typically will be below about 3 times 10.sup.7 cell/ml.
  • the bioactive material in the invention is almost exclusively from platelets.
  • the range of the mean platelet volume of the platelets being sequestered is in the range of about 6.6 to 8.4 femtoliters, with an average of about 7.7 femtoliters; this may indicate that the platelets being sequestered are relatively larger or younger than the overall population of platelets.
  • Activation of growth factors may occur in a variety of manners, by a variety of substances known as activators or agonists.
  • said activation results from lysine and the inclusion of an activator or agonist selected from the group consisting of thrombin, glass, collagen, serotonin, adenosine diphosphate (ADP) and acetylcholine (ACH), and combinations thereof.
  • said growth factors are included within concentrated platelets, and said activation results from the inclusion of thrombin.
  • ADP adenosine diphosphate
  • ACH acetylcholine
  • the activator or agonist added to the platelet and plasma concentrate is in an amount sufficient to facilitate the formation of the coagulum (gel) having a predetermined viscosity while sufficiently activating growth factors present in the composition.
  • the amount of thrombin generally ranges between about 100 U and about 10,000 U, preferably about 900 LJ and about 1100 U, most preferably about 1000 U per 10 cc of platelet concentrate.
  • Thrombin is available as Thrombogen.RTM. thrombin, topical USP (bovine origin) in vials containing 5000 units thrombin (Johnson & Johnson Medical Inc., Arlington, Tex., USA).
  • the invention described herein may also include a method of treating a wound, comprising the steps of applying a sufficient amount of a composition of matter comprising growth factors to enhance healing of the wound.
  • Said method of treating a wound may include the use of any of the compositions described herein; it may also include the use of any composition made by any of the methods described herein.
  • the composition may remain on the wound for as long as 5 days, and perhaps longer depending upon the circumstances such as the location of the wound and other wound characteristics.
  • the composition and method described herein are especially useful for the treatment of chronic wounds, they may also be useful in the treatment of acute wounds.
  • Pooled or single donor platelets (containing at least 5 x 10 to the 9 th platelets) about 40ml to 50ml per bag.
  • Thaw the fresh frozen plasma ad insert an amount into the platelet plug container as to cause a platelet count of between 250,000 to 4,500,000 platelets per milliliter. Being very careful that the fresh plasma does not stay thawed for more than 4 hours. Gently rotate back and forth to cause the platelets and plasma to mix well. Pipette the desired amount of PRP into the vials to be lyophilized and place the stoppers in place.
  • the first cycle should be for 48 hours.
  • the second cycle should be for 12 hours.
  • Thrombin (bovine) 1000u per milliliter Place on the wound and cover with a moist gauze. Cover the wound with an exclusive dressing. Allow too stay in place for 4 to 7 days without changing the dressing.
  • Pooled or single donor platelets (containing at least 5 x 10 to the 9 th platelets) about 40ml to 50ml per bag.
  • GHK-Cu ranging from .02mg/ml to .5mg/ml in liquid form Method: Remove the platelets from the bag and place in a 50ml tube and centrifuge for 15-20 min. at ⁇ OOOrpms to form a platelet plug, which is known in the art.
  • the first cycle should be for 48 hours.
  • the second cycle should be for 12 hours.
  • 10ml of the PRP was Lyophilized and capped.
  • Pooled or single donor platelets (containing at least 5 x 10 to the 9 th platelets) about 40ml to 50ml per bag.
  • GHK-Cu ranging from .02mg/ml to .5mg/ml gauze Method:
  • the first cycle should be for 48 hours.
  • the second cycle should be for 12 hours.
  • 10ml of the PRP was Lyophilized and capped.
  • 10ml vials of LPRP was Lyophilized and capped.
  • Pooled or single donor platelets (containing at least 5 x 10 to the 9 th platelets) about 40ml to 50ml per bag.
  • the first cycle should be for 48 hours.
  • the second cycle should be for 12 hours.
  • 10ml of the PRP was Lyophilized and capped.
  • the Fifth Method for Human blood components obtained from a blood bank or collection center The components needed: Pooled or single donor platelets (containing at least 5 x 10 to the 9 th platelets) about 40ml to 50ml per bag.
  • Vitamin C is an amount equal to 10%
  • the first cycle should be for 48 hours.
  • the second cycle should be for 12 hours.
  • 10ml of the PRP was Lyophilized and capped.
  • Vitamin C A.K.A. (Ascorbic Acid) Place on the wound and cover with a moist gauze. Cover the wound with an exclusive dressing. Allow too stay in place for 4 to 7 days without changing the dressing.
  • the sixth Method for Human blood components obtained from a blood bank or collection center The components needed: Pooled or single donor platelets (containing at least 5 x 10 to the 9 th platelets) about 40ml to 50ml per bag.
  • Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms.
  • Vitamin C is an amount equal to 10%
  • GHK-Cu ranging from .02mg/ml to .5mg/ml in liquid form
  • the first cycle should be for 48 hours.
  • the second cycle should be for 12 hours.
  • Single donor platelets obtained from apheresis about 40ml to 50ml per bag. Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms. 50ml tubes. Pipettes from .01 ul to 50ml. Vials with stoppers and caps. Method:
  • the first cycle should be for 48 hours.
  • the second cycle should be for 12 hours.
  • Pooled or single donor platelets (containing at least 5 x 10 to the 9 th platelets) about 40ml to 50ml per bag. Pooled or single donor fresh frozen plasma (about 250ml) per bag. Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms.
  • the first cycle should be for 48 hours.
  • the second cycle should be for 12 hours.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Veterinary Medicine (AREA)
  • Chemical & Material Sciences (AREA)
  • Public Health (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • General Health & Medical Sciences (AREA)
  • Epidemiology (AREA)
  • Engineering & Computer Science (AREA)
  • Immunology (AREA)
  • Hematology (AREA)
  • Cell Biology (AREA)
  • Zoology (AREA)
  • Biomedical Technology (AREA)
  • Molecular Biology (AREA)
  • Inorganic Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Biotechnology (AREA)
  • Developmental Biology & Embryology (AREA)
  • Virology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Medicines Containing Material From Animals Or Micro-Organisms (AREA)
  • Materials For Medical Uses (AREA)

Abstract

This invention relates to an improved lyophilized platelet rich plasma used to make a platelet gel wound healant, and methods of preparation and use thereof for healing wounds are disclosed. the improved wound healant comprises therapeutically effective amounts of activated growth factors, platelet ghost, plasma (know as the plasma back bone), white blood cells with optional none, one or more additional anti-oxidant such as vitamin A and/or c and/or E, and/or none one or more antibiotics and/or GHK-Cu.

Description

LYOPHILIZED PLATELET RICH PLASMA FOR THE USE IN WOUND HEALING AND BONE OR TISSUE GRAFTS OR REPAIR
BACKGROUND OF THE INVENTION 1) Field of the Invention
The invention relates to an improved Lyophilized platelet rich plasma used to make a platelet gel wound healant, and methods of preparation and use thereof for healing wounds are disclosed. The improved wound healant comprises a therapeutically effective amount of activated growth factors with optional none, one or more additional anti-oxidant such as vitamin A and/or C and/or E, and/or one or more antibiotics and/or GHK-Cu (produced by
ProCyte Inc.).
The present invention relates to improved Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts and methods of making and use thereof of Lyophilized or fixed platelets.
The present invention generally relates to the therapeutic uses of blood platelets and fresh plasma that has been lyophilized, and more particularly to manipulations or modifications of platelets and plasma, such as in preparing freeze-dried compositions that can be rehydrated at the time of application and which when rehydrated have a normal response to thrombin and other agonists with respect to that of fresh platelets. The inventive compositions are useful in applications such as wound care. 2) Description of Related Art
Several techniques for preservation of platelets have been developed over the past few decades. Cryopreservation of platelets using various agents, such as glycerol (Valeri et al., Blood, 43, 131-136, 1974) or dimethyl sulfoxide, "DMSO" (Bock et al., Transfusion, 35, 921-924, 1995), as the cryoprotectant has been done with some success. The best results have been obtained with DMSO. However, a considerable fraction of these cells are partly lysed after thawing and have the shape of a balloon. These balloon cells are not responsive to various agonists, so that overall responsiveness of frozen thawed platelets to various agonists is reduced to less than 35% compared with fresh platelets. The shelf life of cryopreserved DMSO platelets at -δO.degree. C. is reported to be one year, but requires extensive washing and processing to remove cryoprotective agents, and even then the final product has a severe reduction in ability to form a clot. Attempts to dry platelets by Lyophilization have been described with paraformaldehyde fixed platelets (Read et al., Proc. Natl. Acad Sci. USA, 92, 397-401 , 1995). U.S. Pat. No. 5,902,608, issued May 11 , 1999, inventors Read et al. describe and claim a surgical aid comprising a substrate on which fixed, dried blood platelets are carried. These dried blood platelets are fixed by contacting the platelets to a fixative such as formaldehyde, paraformaldehyde, gutaraldehyde, or permanganate. Proper functioning of lyophilized platelets that have been fixed by such fixative agents in hemostasis is questionable.
Spargo et al., U.S. Pat. No. 5,736,313, issued Apr. 7, 1998, has described a method in which platelets are loaded overnight with an agent, preferably glucose, and subsequently lyophilized. The platelets are preincubated in a preincubation buffer and then are loaded with carbohydrate, preferably glucose, having a concentration in the range of about 100 mM to about 1.5M. The incubation is taught to be conducted at about 10.degree. C. to about 37.degree. C, most preferably about 25.degree. C. U.S. Pat. No. 5,827,741 , Beattie et al., issued Oct. 27, 1998, discloses cryoprotectants for human platelets, such as dimethylsulfoxide and trehalose.
The platelets may be suspended, for example, in a solution containing a cryoprotectant at a temperature of about 22.degree. C. and then cooled to below 15. degree. C. This incorporates some cryoprotectant into the cells.
Other workers have sought to load platelets with trehalose through use of electroporation before drying under vacuum. However, electroporation is very damaging to the cell membranes and is believed to activate the platelets. Activated platelets have dubious clinical value. Accordingly, a need exists for the effective and efficient preservation of platelets such that they maintain, or preserve, their biological properties, particularly their response to platelet agonists such as thrombin, and which they release their growth factors. Further, it would also be useful to expand the types of present vehicles that are useful for wound care. There have been many different substances and methods developed in the past for treating wounds, depending upon the type and location and severity of the wound. A wound is generally defined as an injury to an area of the body of a human or animal. Although injury to the surface of the skin is the most well known type of wound, the surfaces of internal organs may also be wounded, such as during surgery, rupture of the spleen or liver, or resulting from traumatic blows to the body surface in the vicinity of an internal organ.
Medical practice characterizes wounds as chronic or acute, according to the persistence and severity of the wound. A chronic wound is one that is prolonged or lingering, rather than promptly healed. An acute wound is one that occurs relatively quickly, and heals relatively quickly as well. Tissue wounds may have a wide spectrum of manifestations, as small as merely an abnormal microscopic tear or fissure in tissue (or a surface thereof), or as large as the abrasion or ablation of the skin covering a substantial portion of the body, such as in a burn victim. Acute wounds covering a large or movable surface are usually the most difficult to guard from infection, and to heal.
Blood and bodily fluids include various substances that affect wound healing. The blood is the primary medium for delivering healing agents to the wound site, and for transporting foreign or harmful substances away from the wound. Whole blood is primarily comprised of three main types of cells suspended in a protein rich solution known as plasma. The three main cell types in whole blood are erythrocytes (a.k.a. red blood cells), leukocytes (a.k.a. white blood cells) and thrombocytes (a.k.a. platelets). The red blood cells are the iron-containing cells that facilitate the transport and transfer of oxygen to body tissue, and the removal of carbon dioxide. The white blood cells perform a variety of functions such as phagocytosis of foreign bodies and production of antibodies, and are primarily responsible for fighting infection and foreign substances within the blood or wound site. Platelets perform many functions such as plugging leaks in blood vessels and helping begin the process leading to the formation of a blood clot; platelets contain substances known as growth factors that facilitate the formation of new tissue.
Although there are several methods for separating whole blood into its various components, one of the most convenient and expeditious methods is accomplished by differentially centrifuging blood or some of its components
(i.e., apheresis). Using apheresis, the red and white blood cells and plasma may be separated out and returned to the donor's or patient's body, leaving the sequestered platelets in essentially concentrated form for use in wound healing techniques. This may be preformed at a blood collection center, blood bank, at the Physicians Clinic or Hospital. From blood extracted from a patient or from the other forms of collection methods, the platelets may thus be obtained and activated for use on the patient; methods of using a patient's own blood are called "autologous" or "autogenic" donor methods. Another method using blood donated by one or more third parties for use by a patient are called "homologous" or "heterologous" donor methods, or collectively called "allogenic" methods. One of the proteins suspended in plasma is fibrinogen, which reacts with substances released into (or attracted by) wound sites to produce sticky strands of fibrin. Such reactions result in the cross linking of the fibrin strands to form a mesh that holds and supports the deposit or growth of other tissue materials at the wound site. Therefore, the need for fresh plasma also known in the art "The Plasma Back Bone".
The wound healing process is generally considered to occur in several stages, generally known as the healing cascade. After tissue injury, platelets are among the first cells to appear in the vicinity of the wound. Activation of a platelet by an agonist such as thrombin, or other agonists such as those listed elsewhere herein, leads to the release of granule material from within the platelet. Such granulation activation results in the release of proteins known as growth factors, primarily concentrated in the alpha granules of platelets. These released growth factors stimulate the formation of new tissue; when applied to wounds, growth factors have been known to increase the rate of collagen laydown, vascular ingrowth, fibroblast proliferation and overall healing. The release of a protein known as platelet-derived growth factor (PDGF) is a chemotactic signal for monocytes, neutrophils and fibroblasts, which then move into the wound, to begin the inflammatory stage of the healing process. During this time, monocytes secrete a number of factors including PDGF and transforming growth factor-.beta.i (also found in platelets), which recruits and activates fibroblasts, to begin the repair stage of the healing process. Subsequently, wound healing continues through the process of collagen remodeling within the wound. Thus the importance of the ability of the Lyophilized platelet rich plasma to release their growth factors. Based upon the foregoing general scientific principles, already known in the field are wound sealants made from biological materials obtained primarily from tissue other than blood platelets. An example is wound sealants such as "fibrin glue", which often are essentially a mixture of co-coagulants (thrombin and calcium), concentrated fibrinogen and other coagulation proteins. In most applications, the primary roles of fibrin glue are to seal wound surfaces to prevent loss of blood and other body fluids after surgery, and to provide adhesion between adjacent tissue surfaces. These products form a hard, cast-like covering over the area to be sealed, and tend to be non- yielding to limb movement. While there has been much research concerning fibrin glue, this material belongs to a separate field from the present invention, primarily because fibrin glues typically contain cryoprecipitated proteins without platelets. The use of fibrin glue is discussed extensively in the scientific literature; for example, see the references cited in U.S. Pat. No. 5,585,007 issued to Antanavich et al on Dec. 17, 1996. Wound treatment compositions derived from platelet enriched concentrates are known and possess certain advantages over materials without platelets such as fibrin glue. One reason is that natural wound healing agents are released by the platelets. Further, the concentration of platelets likewise allows for a concentrated amount of wound healing factors. Representative examples of platelet derived wound treatment compositions are described for instance in Hood U.S. Pat. No. 5,733,545; Knighton U.S. Pat. No. 5,165,938; and Gordiner U.S. Pat. No. 5,599,558. This form of treatment has major problems, like time consuming and problems collecting blood sometimes with expensive machines and disposables.
Platelet concentrates are typically isolated by the process of differential centrifugation, which essentially allows separating the blood into at least three different components: packed erythrocytes (red blood cells), plasma and platelet concentrate. Platelet concentrate can be combined with a solution of either sodium or calcium mixed with thrombin ("calcified thrombin"), which instantaneously form a composition of activated platelets that, when made with the necessary viscosity, can be utilized as a wound sealant. The chemical reactions and cascades that normally occur when thrombin is added to the concentrated platelets are indeed complex. See, for instance, Reeder, et al, in Proceedings of the American Academy of Cardiovascular Perfusion, Vol. 14, January 1993. Such wound sealants typically set up into a hard mass covering the application site, thereby sealing the site against further blood loss and external contaminants.
There are a number of disadvantages associated with conventional wound compositions derived from platelet concentrates. For instance, activation of platelets leads to instantaneous hardening of the material and thus requires the physician to both activate and apply the platelet composition to the wound site within seconds of activation. Also, certain platelet compositions must be applied to the wound site on a daily basis and thus require regular blood withdrawal from the patient. This is time consuming and if more of the compositions is needed, then another needle stick is needed and the process time required to make the composition.
Accordingly, an improved Lyophilized platelet enriched plasma wound treatment composition which avoids or diminishes the problems associated with typical platelet enriched wound compositions and would be desirable.
SUMMARY OF THE INVENTION
In one aspect of the present invention, a dehydrated composition is provided comprising freeze-dried platelets and fresh plasma that are effectively to preserve biological properties during freeze-drying and rehydration. These platelets are rehydratable so as to have a normal response to at least one agonist, such as thrombin. For example, substantially all freeze-dried platelets of the invention when rehydrated and mixed with thrombin (1 U/ml) form a clot within three minutes at 37.degree. C. The dehydrated composition can include one or more other agents, such as antibiotics, antifungals, growth factors, or the like, depending upon the desired therapeutic application.
The present invention relates to an improved Lyophilized platelet and plasma composition for wound treatment, a method of making and use thereof. The composition comprises Lyophilized platelets and plasma composition in liquid form to the wound site and will gel to prevent the material from flowing away from the site. Optional antibiotics may be included in the improved composition to prevent infections at the wound site. The presence of the anti-oxidant, including vitamins and non-vitamin anti-oxidants, and other healing promotion materials that do not detract from, substantially interfere with, or even destroy the different thrombin activation reactions. The inventive
Lyophilized platelet gels containing fresh plasma is prepared for topical application at the wound site and avoid the requirement for daily reapplication.
Methods of making and using inventive embodiments are also described. One such method is a process of preparing a dehydrated composition comprising providing a source of platelets and fresh plasma and lyophilizing the platelets and plasma. The rehydration preferably done with de- ionized water.
While the inventive composition is preferably used for topical application to the exterior surface of the chronic wounds such as ulcers of the feet of diabetics, the composition may be applied to facilitate the healing of other wounds such as acute wounds such as surgery burns. However, the composition of matter and the methods described herein are not limited solely to topical application.
The inventive composition increases the amount of growth factors in the wound, and thereby facilitates the promotion of the healing rate. This may be especially important in "wounded" patients, especially those with chronic wounds who may lack sufficient circulation to facilitate the healing cascade. The invention described herein also facilitates the covering of the wound area with a substance that prevents or helps to reduce infection caused by most bacteria. Practice of the invention permits the manipulation or modification of platelets and plasma while maintaining, or preserving, biological properties, such as a response to thrombin. The inventive freeze-dried platelets and plasma including the freeze-dried platelets, are substantially shelf stable at ambient temperatures when packaged in moisture barrier materials.
In most general terms, the invention described herein expands the uses for concentrated platelet materials, especially those in gel form, by improving the speed and convenience of making the composition; the invention described herein also improves the performance of the concentrated platelet composition, by making it more useable for applications over longer periods of time, and by enhancing the wound healing and infection fighting properties.
Another aspect of the present invention involves adding one or more antibiotic substance at one or more times during the processing period so that the resulting concentrated platelet and plasma composition contains either one or a variety of the antibiotics. The use of an antibiotic in concentrated platelet and plasma compositions that enhances the complex healing cascade is indeed novel. The invention disclosed herein involves adding such substances in a manner that does not detract from, substantially interfere with, or even destroy these different reactions, pH balances and potency.
Another aspect of the present invention involves adding one or more vitamins, to the concentrated platelet gel. Vitamins are known to have wound healing and anti-oxidant properties. Representative examples of suitable, but none limiting, vitamins include vitamin E, vitamin A, vitamin C and other retinoids. In yet another aspect of the invention, non-vitamin anti-oxidants may be included in the concentrated platelet gel. Non-limiting representative examples of such anti-oxidants include beta-carotene.
DETAILED DESCRIPTION OF THE INVENTION For the sake of simplicity and to give the claims of this patent application the broadest interpretation and construction possible, the following definitions will apply:
(a) The phrase blood collecting or blood extraction (or similar phrase) includes techniques, materials and apparatus known in the field, such as (for example) inclusion of anticoagulation materials, the use of blood drawing and infusion apparatus. (b) the phrase growth factor means any material(s) promoting growth of a tissue.
(c) The term thrombin may include calcified thrombin, in particular, about 5,000 units of thrombin per 5 ml of 10% of aqueous calcium chloride solution; it may include calcified bovine thrombin as well as autologous thrombin, allogeneic thrombin or recombinant human thrombin.
(d) The term viscosity means those characteristics of the specified material(s) determining the degree of gelation, such as (for example) the firmness or hardness of the material, or the degree to which the material resists flowing like a fluid. (Θ) The term therapeutically effective amount means the amount or amounts of the constituent elements or combination thereof necessary to enhance wound healing such as, for example, the reduction in the volume or surface area of a wound, the increase in the amount of granulation tissue or other biological material facilitating collagen laydown, vascular ingrowth, fibroblast proliferation or overall healing; all of the versions of the invention described herein are assumed to have the therapeutically effect amount(s) of constituent substances, or combinations thereof.
(f) the term anti-oxidant refers to any material(s) having anti-oxidant properties. Anti-oxidant would include, without limitation, vitamins such as vitamins C, A and E and non-vitamins such as -carotene.
Also for the sake of simplicity, the conjunctive "and" may also be taken to include the disjunctive "or", and vice versa, whenever necessary to give the claims of this patent application the broadest interpretation and construction possible. Likewise, when the plural form is used it may be taken to include the singular form and vice versa.
In most general terms, the invention includes a wound healant composition comprising activated growth factors and ascorbic acid. In the prevalent version of the invention, said growth factors are included within platelets. The body produces many substances generally known as growth factors, and these growth factors are contemplated for use in the present invention. The preferred growth factors for use in the present invention are selected from the group consisting of platelet-derived growth factor (PDGF), platelet-derived angiogenesis factor (PDAF), vascular endotheial growth factor (VEGF), platelet-derived epidermal growth factor (PDEGF), platelet factor 4 (PF-4), transforming growth factor .beta. (TGF-B), acidic fibroblast growth factor (FGF-A), basic fibroblast growth factor (FGF-B), transforming growth factor .alpha. (TGF-A), insulin-like growth factors 1 and 2 (IGF-1 and IGF-2), .beta, thromboglobulin-related proteins (BTG), thrombospondin (TSP), fibronectin, von Wallinbrand's factor (vWF), fibropeptide A, fibrinogen, albumin, plasminogen activator inhibitor 1 (PAI-1 ), osteonectin, regulated upon activation normal T cell expressed and presumably secreted (RANTES), gro-.alpha., vitronectin, fibrin D-dimer, factor V, antithrombin III, immunoglobulin-G (IgG), immunoglobulin-M (IgM), immunoglobulin-A (IgA), a2-macroglobulin, angiogenin, Fg-D, elastase, keratinocyte growth factor (KGF), epidermal growth factor (EGF), fibroblast growth factor (FGF), tumor necrosis factor (TNF), fibroblast growth factor (FGF) and interleukin-1 (IL-1 ), Keratinocyte Growth Factor-2 (KGF-2). and combinations thereof. One of the important characteristics common to each substance, supporting the inclusion of each in this particular group, is that each such substance is known or believed to enhance cell or tissue growth. Moreover, said substances, or various combinations thereof, are known or believed to function together in an unexpected synergistic manner to promote wound healing. Suitable, non- limiting, anti-oxidants useful in the invention include but are not limited to vitamins such as vitamin C (ascorbic acid), vitamin E, vitamin A and other retinoids; and the carotenes such as beta-carotene. In practicing this invention, ascorbic acid as anti-oxidant is particularly preferred.
The platelets are separated from the red blood cells and white blood cells of whole blood, primarily through differential centrifugation, although any suitable method for separating platelets from whole blood may be employed in practicing this invention. The overall composition of the invention disclosed herein may contain incidental amounts of white blood cells, due to the fact that the platelets are rarely totally isolated from the other blood components. It is believed that the present invention contains only minimal or trace amounts of white blood cells; it is believed that the white blood cell count of the present invention typically will be below about 3 times 10.sup.7 cell/ml. The bioactive material in the invention is almost exclusively from platelets. The range of the mean platelet volume of the platelets being sequestered is in the range of about 6.6 to 8.4 femtoliters, with an average of about 7.7 femtoliters; this may indicate that the platelets being sequestered are relatively larger or younger than the overall population of platelets.
Activation of growth factors may occur in a variety of manners, by a variety of substances known as activators or agonists. In the invention described herein, said activation results from lysine and the inclusion of an activator or agonist selected from the group consisting of thrombin, glass, collagen, serotonin, adenosine diphosphate (ADP) and acetylcholine (ACH), and combinations thereof. In a particular and preferred version of the invention, said growth factors are included within concentrated platelets, and said activation results from the inclusion of thrombin. One of the important characteristics common to each substance, supporting the inclusion of each in this particular group, is that each such substance is known or believed to enhance cell or tissue growth in addition to the ability to activate platelets. Moreover, said substances, or various combinations thereof, are known or believed to function together in an unexpected synergistic manner to promote wound healing. The activator or agonist added to the platelet and plasma concentrate is in an amount sufficient to facilitate the formation of the coagulum (gel) having a predetermined viscosity while sufficiently activating growth factors present in the composition. In the preferred case where thrombin is employed as the activator to produce a final soft gel wound composition in a soft gel form, the amount of thrombin generally ranges between about 100 U and about 10,000 U, preferably about 900 LJ and about 1100 U, most preferably about 1000 U per 10 cc of platelet concentrate. Thrombin is available as Thrombogen.RTM. thrombin, topical USP (bovine origin) in vials containing 5000 units thrombin (Johnson & Johnson Medical Inc., Arlington, Tex., USA).
Since the admixture of thrombin or other agonists will activate growth factors, the thrombin (or other agonists/activators) should usually be the last substance to be mixed immediately before it is desired that the gelatinous state be set up. Besides a method of making a wound healant composition, the invention described herein may also include a method of treating a wound, comprising the steps of applying a sufficient amount of a composition of matter comprising growth factors to enhance healing of the wound. Said method of treating a wound may include the use of any of the compositions described herein; it may also include the use of any composition made by any of the methods described herein.
Once applied to a wound, the composition may remain on the wound for as long as 5 days, and perhaps longer depending upon the circumstances such as the location of the wound and other wound characteristics. Although the composition and method described herein are especially useful for the treatment of chronic wounds, they may also be useful in the treatment of acute wounds.
EXAMPLE 1 :
Preparation of Lyophilized Platelet Rich Plasma for the use in Wound Healing (Chronic or Acute) and Bone or Tissue Grafts.
(a) The First Method for Human blood components obtained from a
blood bank or collection center: The components needed:
Pooled or single donor platelets (containing at least 5 x 10 to the 9th platelets) about 40ml to 50ml per bag.
Pooled or single donor fresh frozen plasma (about 250ml) per bag. Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms. 50ml tubes.
Pipettes from .01 ul to 50ml Vials with stoppers and caps.
Method:
Remove the platelets from the bag and place in a 50ml tube and centrifuge for 15-20 min. at 5000rpms to form a platelet plug, which is known in the art. Remove and discard the platelet poor plasma from the tube of platelets.
Thaw the fresh frozen plasma ad insert an amount into the platelet plug container as to cause a platelet count of between 250,000 to 4,500,000 platelets per milliliter. Being very careful that the fresh plasma does not stay thawed for more than 4 hours. Gently rotate back and forth to cause the platelets and plasma to mix well. Pipette the desired amount of PRP into the vials to be lyophilized and place the stoppers in place.
Lyophilize at once (take care as to not allow any warming to occur). The first cycle should be for 48 hours. The second cycle should be for 12 hours.
For this example 10ml of the PRP was Lyophilized and capped. To apply to the wound using the 10ml vials of LPRP:
Rehydrate using (10ml) de-ionized water and allow to stand for at least 10 to 15 minutes. Remove the LPRP using a 20ml syringe.
Add 1ml of Thrombin (bovine) 1000u per milliliter Place on the wound and cover with a moist gauze. Cover the wound with an exclusive dressing. Allow too stay in place for 4 to 7 days without changing the dressing.
Repeat if necessary. EXAMPLE 2:
(a) The Second Method for Human blood components obtained from a blood bank or collection center: The components needed:
Pooled or single donor platelets (containing at least 5 x 10 to the 9th platelets) about 40ml to 50ml per bag.
Pooled or single donor fresh frozen plasma (about 250ml) per bag. Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms. 50ml tubes. Pipettes from .01 ul to 50ml Vials with stoppers and caps.
GHK-Cu ranging from .02mg/ml to .5mg/ml in liquid form Method: Remove the platelets from the bag and place in a 50ml tube and centrifuge for 15-20 min. at δOOOrpms to form a platelet plug, which is known in the art.
Remove and discard the platelet poor plasma from the tube of platelets. Thaw the fresh frozen plasma ad insert an amount into the platelet plug container as to cause a platelet count of between 250,000 to 4,500,000 platelets per milliliter. Being very careful that the fresh plasma does not stay thawed for more than 4 hours. Add an amount of GHK-Cu to equal 1ml per 9ml of LPRP Gently rotate back and forth to cause the platelets and plasma to mix well. Pipette the desired amount of PRP into the vials to be lyophilized and place the stoppers in place.
Lyophilize at once (take care as to not allow any warming to occur). The first cycle should be for 48 hours. The second cycle should be for 12 hours. For this example 10ml of the PRP was Lyophilized and capped. To apply to the wound using the 10ml vials of LPRP:
Re-hydrate using (10ml) de-ionized water and allow to stand for at least 10 to 15 minutes. Remove the LPRP using a 20ml syringe. Add 1 ml of Thrombin (bovine) 100Ou per milliliter Place on the wound and cover with a moist gauze. Cover the wound with an exclusive dressing. Allow too stay in place for 4 to 7 days without changing the
dressing. Repeat if necessary.
(b) The Third Method for Human blood components obtained from a blood bank or collection center: The components needed:
Pooled or single donor platelets (containing at least 5 x 10 to the 9th platelets) about 40ml to 50ml per bag.
Pooled or single donor fresh frozen plasma (about 250ml) per bag. Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms. 50ml tubes.
Pipettes from .01 ul to 50ml Vials with stoppers and caps.
GHK-Cu ranging from .02mg/ml to .5mg/ml gauze Method:
Remove the platelets from the bag and place in a 50ml tube and centrifuge for 15-20 min. at 5000rpms to form a platelet plug, which is known in the art.
Remove and discard the platelet poor plasma from the tube of platelets. Thaw the fresh frozen plasma ad insert an amount into the platelet plug container as to cause a platelet count of between 250,000 to 4,500,000 platelets per milliliter. Being very careful that the fresh plasma does not stay thawed for more than 4 hours. Gently rotate back and forth to cause the platelets and plasma to mix well. Pipette the desired amount of PRP into the vials to be lyophilized and place the stoppers in place.
Lyophilize at once (take care as to not allow any warming to occur). The first cycle should be for 48 hours.
The second cycle should be for 12 hours. For this example 10ml of the PRP was Lyophilized and capped. To apply to the wound using the 10ml vials of LPRP:
Re-hydrate using (10ml) de-ionized water and allow to stand for at least 10 to 15 minutes.
Remove the LPRP using a 20ml syringe. Add 1 ml of Thrombin (bovine) 1000u per milliliter Place on the wound and cover with GHK-Cu gauze. Cover the wound with an exclusive dressing. Allow too stay in place for 4 to 7 days without changing the dressing.
Repeat if necessary.
(c) The Forth Method for Human blood components obtained from a blood bank or collection center: The components needed:
Pooled or single donor platelets (containing at least 5 x 10 to the 9th platelets) about 40ml to 50ml per bag.
Pooled or single donor fresh frozen plasma (about 250ml) per bag. Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms. 50ml tubes. Pipettes from .01 ul to 50ml Vials with stoppers and caps. Vitamin C is an amount equal to 10% Method: Remove the platelets from the bag and place in a 50ml tube and centrifuge for 15-20 min. at 5000rpms to form a platelet plug, which is known in the art.
Remove and discard the platelet poor plasma from the tube of platelets. Thaw the fresh frozen plasma ad insert an amount into the platelet plug container as to cause a platelet count of between 250,000 to 4,500,000 platelets per milliliter. Being very careful that the fresh plasma does not stay thawed for more than 4 hours. Add an amount of Vitamin C to equal 1 ml per 9ml of LPRP Gently rotate back and forth to cause the platelets and plasma to mix well. Pipette the desired amount of PRP into the vials to be lyophilized and place the stoppers in place.
Lyophilize at once (take care as to not allow any warming to occur). The first cycle should be for 48 hours. The second cycle should be for 12 hours. For this example 10ml of the PRP was Lyophilized and capped. To apply to the wound using the 10ml vials of LPRP:
Re-hydrate using (10ml) de-ionized water and allow to stand for at least 10 to 15 minutes. Remove the LPRP using a 20ml syringe. Add 1 ml of Thrombin (bovine) 100Ou per milliliter Place on the wound and cover with a moist gauze. Cover the wound with an exclusive dressing. Allow too stay in place for 4 to 7 days without changing the dressing. Repeat if necessary.
(d) The Fifth Method for Human blood components obtained from a blood bank or collection center: The components needed: Pooled or single donor platelets (containing at least 5 x 10 to the 9th platelets) about 40ml to 50ml per bag.
Pooled or single donor fresh frozen plasma (about 250ml) per bag. Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms. 50ml tubes. Pipettes from .01 ul to 50ml
Vials with stoppers and caps. Vitamin C is an amount equal to 10% Method:
Remove the platelets from the bag and place in a 50ml tube and centrifuge for 15-20 min. at 5000rpms to form a platelet plug, which is known in the art.
Remove and discard the platelet poor plasma from the tube of platelets.
Thaw the fresh frozen plasma ad insert an amount into the platelet plug container as to cause a platelet count of between 250,000 to 4,500,000 platelets per milliliter. Being very careful that the fresh plasma does not stay thawed for more than 4 hours.
Gently rotate back and forth to cause the platelets and plasma to mix well. Pipette the desired amount of PRP into the vials to be lyophilized and place the stoppers in place.
Lyophilize at once (take care as to not allow any warming to occur). The first cycle should be for 48 hours. The second cycle should be for 12 hours. For this example 10ml of the PRP was Lyophilized and capped. To apply to the wound using the 10ml vials of LPRP:
Re-hydrate using (10ml) de-ionized water and allow to stand for at least 10 to 15 minutes.
Remove the LPRP using a 20ml syringe. Add 1 ml of Thrombin (bovine) 100Ou per milliliter
Add 1 ml of 10% Vitamin C, A.K.A. (Ascorbic Acid) Place on the wound and cover with a moist gauze. Cover the wound with an exclusive dressing. Allow too stay in place for 4 to 7 days without changing the dressing.
Repeat if necessary.
(e) The sixth Method for Human blood components obtained from a blood bank or collection center: The components needed: Pooled or single donor platelets (containing at least 5 x 10 to the 9th platelets) about 40ml to 50ml per bag.
Pooled or single donor fresh frozen plasma (about 250ml) per bag.
Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms.
50ml tubes.
Pipettes from .01 ul to 50ml
Vials with stoppers and caps.
Vitamin C is an amount equal to 10%
GHK-Cu ranging from .02mg/ml to .5mg/ml in liquid form
Method:
Remove the platelets from the bag and place in a 50ml tube and centrifuge for 15-20 min. at 5000rpms to form a platelet plug, which is known in the art.
Remove and discard the platelet poor plasma from the tube of platelets. Thaw the fresh frozen plasma ad insert an amount into the platelet plug container as to cause a platelet count of between 250,000 to 4,500,000 platelets per milliliter. Being very careful that the fresh plasma does not stay thawed for more than 4 hours. Add an amount of Vitamin C to equal .05ml to1 ml per 9ml of LPRP Add an amount of GHK-Cu to equal .05ml to1 ml per 9ml of LPRP
Gently rotate back and forth to cause the platelets and plasma to mix well. Pipette the desired amount of PRP into the vials to be lyophilized and place the stoppers in place.
Lyophilize at once (take care as to not allow any warming to occur). The first cycle should be for 48 hours. The second cycle should be for 12 hours.
For this example 10ml of the PRP was Lyophilized and capped. To apply to the wound using the 10ml vials of LPRP:
Re-hydrate using (10ml) de-ionized water and allow to stand for at least 10 to 15 minutes.
Remove the LPRP using a 20ml syringe. Add 1 ml of Thrombin (bovine) 1000u per milliliter Place on the wound and cover with a moist gauze. Cover the wound with an exclusive dressing. Allow too stay in place for 4 to 7 days without changing the dressing.
Repeat if necessary. EXAMPLE 3
(a) The sixth method for human or animal, obtained from apheresis either Autologous or Homologous: The components needed:
Single donor platelets (obtained from apheresis) about 40ml to 50ml per bag. Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms. 50ml tubes. Pipettes from .01 ul to 50ml. Vials with stoppers and caps. Method:
Remove the platelets from the bag of apheresis platelets. Add fresh plasma into the platelets in an amount to cause a platelet count of between 250,000 to 4,500,000 platelets per milliliter. Being very careful that the fresh platelet rich plasma does not stay thawed for more than 6 hours.
Gently rotate back and forth to cause the platelets and plasma to mix well. Pipette the desired amount of PRP into the vials to be lyophilized and place the stoppers in place.
Lyophilize at once (take care as to not allow any warming to occur). The first cycle should be for 48 hours. The second cycle should be for 12 hours.
For this example 10ml of the PRP was Lyophilized and capped. To apply to the wound using the 10ml vials of LPRP: a. Re-hydrate using (10ml) de-ionized water and allow to stand for at least 10 to 15 minutes. b. Remove the LPRP using a 20ml syringe. c. Add 1 ml of Thrombin (bovine) 100Ou per milliliter d. Place on the wound and cover with a moist gauze. e. Cover the wound with an exclusive dressing. f. Allow too stay in place for 4 to 7 days without changing the dressing. g. Repeat if necessary. EXAMPLE 4:
(c) The third method for human blood components obtained from a blood bank and activated prior to Lyophilization with Thrombin (bovine or intrinsic): The components needed:
Pooled or single donor platelets (containing at least 5 x 10 to the 9th platelets) about 40ml to 50ml per bag. Pooled or single donor fresh frozen plasma (about 250ml) per bag. Centrifuge with the capabilities of spinning 50ml tubes up to 5000rpms.
50ml tubes.
Pipettes from .01 ul to 50ml Vials with stoppers and caps.
6. Thrombin and Calcium Chloride or Glass Beads or Glass Wool. Method:
Remove the platelets from the bag and place in a 50ml tube and centrifuge for 15-20 min. at 5000rpms to form a platelet plug, which is known in the art.
Remove and discard the platelet poor plasma from the tube of platelets. Remove 50% of the platelet plug and activate the plug with Thrombin or pass the platelets through either glass wool or glass beads and allow activated platelets to rest for about 15 min. then centrifuge for 15 min. and remove the serum with out the plug. Then pipette equal amounts of the serum into the vials in equal amounts.
Thaw the fresh frozen plasma and insert an amount into the platelet plug container as to cause a platelet count of between 250,000 to 4,500,000 platelets per milliliter. Being very careful that the fresh plasma does not stay thawed for more than 4 hours.
Gently rotate back and forth to cause the platelets and plasma to mix well. Pipette the desired amount of PRP into the vials to be lyophilized and place the stoppers in place.
Lyophilize at once (take care as to not allow any warming to occur). The first cycle should be for 48 hours. The second cycle should be for 12 hours.
For this example 10ml of the PRP was Lyophilized and capped. To apply to the wound using the 10ml vials of ALPRP:
Rehydrate using (10ml) de-ionized water and allow to stand for at least 10 to 15 minutes.
Remove the LPRP using a 20ml syringe. Place on the wound and cover with a moist gauze. Cover the wound with an exclusive dressing. Allow too stay in place for 4 to 7 days without changing the dressing. Repeat if necessary.

Claims

What is claimed:
1. A dehydrated composition, useful for mammalian therapy, comprising: substantially shelf-stable lyophilized platelets selected from the mammalian species for which therapy is intended; said platelets being effectively loaded with plasma to form a platelet and plasma concentrate having a freeze-drying and rehydration capacity, wherein the platelets are rehydratable so as to have a normal response to at least one agonist.
2. A wound healant described in claim 1 , including growth factors included in said lyophilized platelet and plasma concentrate for being activated by lysine and the inclusion of an agonist to said concentrate.
3. A wound healant described in claim 2, wherein said agonist is selected from the group consisting of thrombin, glass, collagen, serotonin, adenosine diphosphate (ADP) and acetylcholine (ACH), and combinations thereof.
4. A wound healant described in claim 1 , wherein said lyophilized platelets are autologous concentrated platelets.
5. A wound healant described in claim 4, including a white blood cell count of below about 3 time 10.sup.7 cells/ml.
6. A wound healant described in claim 1 , wherein said lyophilized platelets are homologous concentrated platelets.
7. A wound healant described in claim 6, including a white blood cell count of below about 3 time 10.sup.7 cells/ml
8. A wound healant composition comprising a therapeutically effective amount of concentrated Lyophilized platelets, plasma and thrombin.
9. A wound healant composition comprising a therapeutically effective amount of concentrated Lyophilized platelets, plasma, at least one anti-oxidant and thrombin.
10. A wound healant composition described in claim 9, wherein said anti-oxidant is selected from the group consisting of retinoids and carotenes.
11. A wound healant composition described in claim 10, wherein said retinoid is selected from the group consisting of vitamin A, vitamin C, and vitamin E.
12. A wound healant composition described in claim 10, wherein said carotene is beta-carotene.
13. A wound healant composition comprising a therapeutically effective amount of concentrated lyophilized platelets, plasma, at least one antibiotic and thrombin.
14. A wound healant described in claim 13, wherein said antibiotic is bacteriocidal to at least Pseudomonas and Klebsella bacteria.
15. A wound healant described in claim 13, wherein said antibiotic is selected from the group consisting of a neosporin, vancomycin and gentamycin, and combinations thereof.
16. A wound healant composition comprising a therapeutically effective amount of concentrated lyophilized platelets, plasma, and GHK-Cu produced by ProCyte Inc. of Seattle, WA
17. A wound healant composition comprising concentrated lyophilized platelets, plasma, at least one retinoid, at least one antibiotic bacteriocidal to at least Pseudomonas and Klebsella, and thrombin.
18. A wound healant composition comprising concentrated platelets in an amount ranging between about 250,000 to about 4,500,000 platelets per milliliter, vitamin C, thrombin in an amount ranging between about 100 U to about 10,000 U per 10 cc of platelet concentrate.
19. A wound healant composition as described in claim 18, including calcium chloride in an amount ranging between about 0.1 mg/ml to about 10 mg/ml.
20. A wound healant composition as described in claim 18, including retinoid selected from the group consisting of vitamin A, vitamin C and vitamin E, and combinations thereof, in effective anti-oxidative amounts.
21. A method of making a wound healant composition, comprising the steps of mixing, in therapeutically effective amount(s), activated lyophilized platelet and plasma concentrate.
22. A method of making a wound healant described in claim 21 , wherein said activated lyophilized platelet and plasma concentrate are obtained from one of a blood bank, collection center, and Apheresis, and thrombin is mixed with said platelets.
23. A method of making a wound healant described in claim 22, said method further comprising, prior to mixing said thrombin, mixing at least one antibiotic in sufficient amount(s) to reduce infection by bacteria.
24. A method of making a wound healant described in claim 21 , said method further comprising, either prior to or after lyophilization, mixing at least one retinoid in sufficient amount(s) to further enhance wound healing.
25. A method of making a wound healant described in claim 21 , wherein said antibiotic is at least bacteriocidal to Pseudomonas and Klebsella bacteria.
26. A method of making a wound healant described in claim 21 , wherein said antibiotic is selected from the group consisting of neosporin, vancomycin and gentamycin, and combinations thereof.
27. A wound healant prepared in accordance with claim 21.
28. A method of making a wound healant, comprising the steps of, prior to or after lyophilization, admixing, in therapeutically effective amount(s), concentrated platelets, ascorbic acid, at least one retinoid, and at least one antibiotic bacteriocidal to at least Pseudomonas and Klebsella bacteria.
29. A wound healant prepared in accordance with claim 28.
30. A method of making a wound healant composition, comprising the steps of: extracting blood from a patient and separating through Apheresis into plasma, red blood cells, and concentrated platelets; removing said plasma; centrifuging said remaining red blood cells and platelets; removing said concentrated platelets; and, mixing, in therapeutically effective amount(s), said concentrated platelets with plasma and thrombin.
31. The method of claim 30, further including the step of lyophilizing said concentrated platelets.
32. A wound healant prepared in accordance with claim 31.
PCT/US2005/031117 2005-08-30 2005-08-30 Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts or repair WO2007027178A2 (en)

Priority Applications (4)

Application Number Priority Date Filing Date Title
PCT/US2005/031117 WO2007027178A2 (en) 2005-08-30 2005-08-30 Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts or repair
CA002621140A CA2621140A1 (en) 2005-08-30 2005-08-30 Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts or repair
EP05793313A EP1943349A2 (en) 2005-08-30 2005-08-30 Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts or repair
BRPI0520502-6A BRPI0520502A2 (en) 2005-08-30 2005-08-30 dehydrated composition usable for mammal therapy, wound healing composition, wound healing method, wound healing, and wound healing method

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
PCT/US2005/031117 WO2007027178A2 (en) 2005-08-30 2005-08-30 Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts or repair

Publications (2)

Publication Number Publication Date
WO2007027178A2 true WO2007027178A2 (en) 2007-03-08
WO2007027178A3 WO2007027178A3 (en) 2007-09-20

Family

ID=37809308

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2005/031117 WO2007027178A2 (en) 2005-08-30 2005-08-30 Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts or repair

Country Status (4)

Country Link
EP (1) EP1943349A2 (en)
BR (1) BRPI0520502A2 (en)
CA (1) CA2621140A1 (en)
WO (1) WO2007027178A2 (en)

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090223080A1 (en) * 2007-03-19 2009-09-10 Hemcon Medical Technologies, Inc. Apparatus and methods for making, storing, and administering freeze-dried materials such as freeze-dried plasma
WO2011124280A1 (en) * 2010-04-08 2011-10-13 INSERM (Institut National de la Santé et de la Recherche Médicale) Zeodration method for the preservation of blood platelets
US8449520B2 (en) 2007-03-19 2013-05-28 HemCon Medical Technologies Inc. Apparatus and methods for making, storing, and administering freeze-dried materials such as freeze-dried plasma
WO2015071364A1 (en) * 2013-11-14 2015-05-21 Nte-Sener, S.A. Method for obtaining a cytokine-rich composition and composition obtained by means of this method

Family Cites Families (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5378461A (en) * 1991-07-12 1995-01-03 Neigut; Stanley J. Composition for the topical treatment of skin damage
WO1998012274A1 (en) * 1996-09-23 1998-03-26 Chandrashekar Pathak Methods and devices for preparing protein concentrates
AU2001234978A1 (en) * 2000-02-10 2001-08-20 The Regents Of The University Of California Therapeutic platelets and methods

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090223080A1 (en) * 2007-03-19 2009-09-10 Hemcon Medical Technologies, Inc. Apparatus and methods for making, storing, and administering freeze-dried materials such as freeze-dried plasma
US8449520B2 (en) 2007-03-19 2013-05-28 HemCon Medical Technologies Inc. Apparatus and methods for making, storing, and administering freeze-dried materials such as freeze-dried plasma
WO2011124280A1 (en) * 2010-04-08 2011-10-13 INSERM (Institut National de la Santé et de la Recherche Médicale) Zeodration method for the preservation of blood platelets
US8802363B2 (en) 2010-04-08 2014-08-12 Inserm (Institut National De La Sante Et De La Recherche Medicale) Zeodration method for the preservation of blood platelets
WO2015071364A1 (en) * 2013-11-14 2015-05-21 Nte-Sener, S.A. Method for obtaining a cytokine-rich composition and composition obtained by means of this method

Also Published As

Publication number Publication date
EP1943349A2 (en) 2008-07-16
CA2621140A1 (en) 2007-03-08
BRPI0520502A2 (en) 2009-06-13
WO2007027178A3 (en) 2007-09-20

Similar Documents

Publication Publication Date Title
US8821858B2 (en) Lyophilized platelet rich plasma for the use in wound healing (chronic or acute) and bone or tissue grafts or repair
US6524568B2 (en) Enriched platelet wound healant
US6303112B1 (en) Enriched platelet wound healant
US8734854B2 (en) Process for removing growth factors from platelets
AU596954B2 (en) Wound healing agents
US7811558B2 (en) Use of stabilized platelets as hemostatic agent
US10111906B2 (en) Serum fraction of platelet-rich fibrin
JPH07507304A (en) Pharmaceutically acceptable fixed dried human blood platelets
US20230248772A1 (en) Products and methods using a platelet-derived hemostatic agent for controlling bleeding and improving healing
AU768543B2 (en) Improved enriched platelet wound healant
EP1943349A2 (en) Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts or repair
US10639333B2 (en) Process for removing growth factors from platelets
MX2008002958A (en) Lyophilized platelet rich plasma for the use in wound healing and bone or tissue grafts or repair
Murdiastuti et al. Effect of freeze-drying process on collagen-activated platelet-rich plasma into platelet derived growth factor-AB level
US20220280606A1 (en) Method of preparing a de-fibrinated platelet lysate, and uses of said method
Nair et al. Blood-derived products in wound healing and repair

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application
ENP Entry into the national phase

Ref document number: 2621140

Country of ref document: CA

WWE Wipo information: entry into national phase

Ref document number: MX/a/2008/002958

Country of ref document: MX

NENP Non-entry into the national phase

Ref country code: DE

WWE Wipo information: entry into national phase

Ref document number: 1277/CHENP/2008

Country of ref document: IN

WWE Wipo information: entry into national phase

Ref document number: 2005793313

Country of ref document: EP

ENP Entry into the national phase

Ref document number: PI0520502

Country of ref document: BR

Kind code of ref document: A2