WO2014027363A1 - A growth factor concentrate for treating hair loss - Google Patents
A growth factor concentrate for treating hair loss Download PDFInfo
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- WO2014027363A1 WO2014027363A1 PCT/IN2013/000268 IN2013000268W WO2014027363A1 WO 2014027363 A1 WO2014027363 A1 WO 2014027363A1 IN 2013000268 W IN2013000268 W IN 2013000268W WO 2014027363 A1 WO2014027363 A1 WO 2014027363A1
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- 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
- A61K38/18—Growth factors; Growth regulators
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- 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
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- 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
- A61K38/18—Growth factors; Growth regulators
- A61K38/1808—Epidermal growth factor [EGF] urogastrone
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- 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
- A61K38/18—Growth factors; Growth regulators
- A61K38/1825—Fibroblast growth factor [FGF]
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- 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
- A61K38/18—Growth factors; Growth regulators
- A61K38/1841—Transforming growth factor [TGF]
-
- 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
- A61K38/18—Growth factors; Growth regulators
- A61K38/1858—Platelet-derived growth factor [PDGF]
-
- 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
- A61K38/18—Growth factors; Growth regulators
- A61K38/1858—Platelet-derived growth factor [PDGF]
- A61K38/1866—Vascular endothelial growth factor [VEGF]
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q7/00—Preparations for affecting hair growth
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
- A61K2800/40—Chemical, physico-chemical or functional or structural properties of particular ingredients
- A61K2800/59—Mixtures
- A61K2800/592—Mixtures of compounds complementing their respective functions
- A61K2800/5922—At least two compounds being classified in the same subclass of A61K8/18
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
- A61K2800/80—Process related aspects concerning the preparation of the cosmetic composition or the storage or application thereof
- A61K2800/84—Products or compounds obtained by lyophilisation, freeze-drying
Definitions
- This invention relates to a growth factor concentrate for treating hair loss.
- Hair loss due to androgenetic alopecia is a common disorder affecting both men and women.
- the incidence of androgenetic alopecia is generally considered to be greater in males than in females, although some evidence suggests that the apparent differences in incidence may be a reflection of different expression in males and females.
- This condition is also known as Male Pattern Hair Loss (MPHL) in men and Female Pattern Hair Loss (FPHL) in women.
- MPHL Male Pattern Hair Loss
- FPHL Female Pattern Hair Loss
- Alopecia in these cases is characterized by thinning of hair as opposed to follicular loss.
- MPHL does not appear until after puberty, usually manifesting in the third decade and in almost all cases by the fourth decade of life.
- MPHL typically begins with bi-temporal recession, followed by progressive thinning in the frontal and vertex areas of the scalp and recession of the frontal hairline is common. Over time, the frontal and vertex thinning areas may merge, resulting in near complete visible hair loss over the top of the scalp.
- FPHL may present as late as the sixth decade of life and is characterized by diffuse thinning in the frontal and parietal areas of the scalp with preservation of the frontal hairline being the norm.
- DHT dihydrotestosterone
- MPHL is caused by a genetic sensitivity of hair follicles to DHT. This hormone causes follicles to shrink or "miniaturize”. In turn, this shortens their lifespan and prevents them from producing hair normally.
- DHT double hexamate
- AGA is a consequence of the anabolic effect of androgens such as hormonal changes leading to structural changes in skin and scalp which in turn cause hair loss.
- minoxidil produced at least moderate cosmetic improvement in only 24% of individuals (Olsen E, Weiner MS, DeLong ER, et al. Topical minoxidil in early male pattern baldness. J Am Acad Dermatol 1985; 13: 185-92).
- minoxidil is also a vasodilator, it is speculated that by widening blood vessels and opening potassium channels, it allows more oxygen, blood and nutrients to reach the follicle. This effect of minoxidil can also cause follicles in the telogen phase to shed, usually soon to be replaced by new, thicker hair (in a new anagen phase).
- minoxidil has severe side effects.
- the alcohol present in topical preparations may dry the scalp, resulting in dandruff.
- Side effects of oral minoxidil may include swelling of the face and extremities, rapid and irregular heartbeat, light headedness, cardiac lesions, and focal necrosis of the papillary muscle and subendocardial areas of the left ventricle.
- Finasteride is a azasteroid and since the drug does not cure the underlying genetic causes of AGA, ceasing daily administration, results in the shedding of regrown hair and the resumption of hair loss. Therefore, continual use of finasteride is necessary . if the cosmetic benefit is to be sustained. Finasteride is usually well tolerated. However, there is a small risk of transient impotence as an adverse effect, which is reversible upon treatment cessation (Amichai B, Grunwald MH, Sobel R. 5a-reductase inhibitors: a new hope in dermatology? Int J Dermatol 1997; 36: 182-4). Zimber et.
- HSC hair stimulating complex
- an intra-dermally, sub-dermally or topically administrable growth factor concentrate derived from approximately 500 l O 6 to 1500 x 10 6 human platelets per ml for treating hair loss, the concentrate comprising approximately 800 to 1200 pg/ml of Epidermal growth factor (EGF), 20 to 80 pg/ml of Vascular Endothelial growth factor (VEGF), 15 to 30 pg/ml of Basic fibroblast growth factor (b-FGF), 30000 to 40000 pg/ml of Transforming growth factor- ⁇ (TGF- ⁇ ) and 100000 to 200000 pg/ml of Platelet Derived growth factor-AB (PDGF-AB).
- EGF Epidermal growth factor
- VEGF Vascular Endothelial growth factor
- b-FGF Basic fibroblast growth factor
- TGF- ⁇ Transforming growth factor- ⁇
- PDGF-AB Platelet Derived growth factor-AB
- a lyophilized, intra-dermally, sub-dermally or topically administrable growth factor concentrate derived from approximately 500 x l O 6 to 1500 x l O 6 human platelets per ml for treating hair loss, the concentrate comprising 800 to 1200 pg/ml of EGF, 20 to 80 pg/ml of VEGF, 15 to 30 pg/ml of b-FGF, 30000 to 40000 pg/ml of TGF- ⁇ and 100000 to 200000 pg/ml of PDGF-AB suspended in an isotonic medium along with excipients for lyophilisation.
- a therapeutic composition for intra-dermal, sub-dermal or topical administration comprising the growth factor concentrate in combination with supplemental constituents including blood, saline, silver nanoparticles, hyaluronic acid, immuno-modulatory peptides, growth factors, hormones, antibiotics, monoclonal antibodies, recombinant receptors, carriers or combinations thereof.
- a method of treatment of hair loss comprising injecting a growth factor concentrate into the scalp first into the subcutaneous fat followed by deep dermal injection and lastly sub-dermal injection, wherein the growth factor concentrate is derived from approximately 875 * 10 6 human platelets per ml for treating hair loss, the concentrate comprising 800 to 1200 pg/ml of Epidermal growth factor (EGF), 20 to 80 pg/ml of VEGF, 15 to 30 pg/ml of b-FGF, 30000 to 40000 pg/ml of TGF- ⁇ and 100000 to 200000 pg/ml of PDGF-AB.
- EGF Epidermal growth factor
- the growth factor concentrate may also be administered by micro-needling or by meso-gun or a combination of the above methods of administration. It is to be understood that both the foregoing general description and the following detailed description of the present embodiments of the invention are intended to provide an overview or framework for understanding the nature and character of the invention as it is claimed. The accompanying graphical representations are included to substantiate the invention and are incorporated into and constitute a part of this specification. BRIEF DESCRIPTION OF THE DRAWINGS
- FIG. lA and FIG IB respectively show a photographic comparison between hair growth before treatment and at two months after administration of the 3 rd dose of growth factor concentrate (GFC), the treatment consisting of three intra-dermal injections of GFC derived from 875 * 10 6 platelets per ml, each subsequent injection being given at an interval of one month from the previous injection.
- GFC growth factor concentrate
- FIG. 2 shows total hair count per sq. cm. of ten patients' scalps using a folliscope before treatment and at two months and at four months after administration of the 3rd dose of GFC, each subsequent injection being given at an interval of one month from the previous injection.
- FIG. 3 shows the caliber of ten patients' hair before treatment and at two months and at four months after administration of the 3rd dose of GFC, each subsequent injection being given at an interval of one month from the previous injection.
- FIG. 4 shows the mean total hair count per sq. cm. of ten patients' scalps using a folliscope before treatment and at two and four months after administration of the 3rd dose of GFC, each subsequent injection being given at an interval of one month from the previous injection.
- FIG. 5 shows the mean caliber of ten patients' hair before treatment and at two and four months after administration of the 3rd dose of GFC, each subsequent injection being given at an interval of one month from the previous injection.
- growth factor concentrate or "GFC” as used in the specification refers to a standardized concentration of growth factors prepared according to an embodiment of the invention, the growtli factors being derived from the cryo-stimulation of platelets which have been counted, the platelets being sourced from human blood.
- the GFC also contains cytokines, chemokines, adhesive proteins and other modulatory peptides.
- multiple electrolyte isotonic solution as used in the specification in the context of the invention comprises sodium chloride, (NaCl); sodium gluconate (C 6 HnNa0 7 ); sodium acetate trihydrate, (C2H 3 Na0 2 » 3H 2 0); potassium chloride, (KC1); and magnesium chloride, (MgCl2'6H 2 0) and human serum albumin.
- each 100 ml of multiple electrolyte isotonic solution contains 526 mg of sodium chloride, (NaCl); 502 mg of sodium gluconate (C 6 HnNa07); 368 mg of sodium acetate trihydrate, (C 2 H 3 Na0 2 *3H 2 0); 37 mg of potassium chloride, (KC1); and 30 mg of magnesium chloride, (Mg0 2 « 6H 2 0) and 1 to 5% of human serum albumin.
- the pH is adjusted with sodium hydroxide to 7.4 (or 6.5 to 8.0), sterile filtered and tested for endotoxin.
- platelet free plasma as used in the specification in the context of the invention comprises the supernatant collected from plasma that has been centrifuged at about 17610 g and then sterile filtered.
- platelet poor plasma or “PPP” as used in the specification in the context of the invention comprises supernatant collected from whole blood that has been centrifuged at about 2720 g.
- baseline as used in the specification in the context of the invention refers to the hair count of a study subject prior to administration of GFC according to an embodiment of the invention.
- the isotonic medium in the growth factor concentrate is multiple electrolyte isotonic solution, plasma, platelet free plasma, platelet poor plasma or a combination thereof.
- the concentrate is derived from approximately 875 x lO 6 human ' platelets per ml.
- the growth factor concentrate according to an embodiment of the invention is used for treatment of hair loss, particularly for treatment of androgenetic alopecia.
- the therapeutic composition according to an embodiment of the invention can be in the form of a cream, gel, aqueous solution, spray-aerosol or transdermal patch.
- three doses of the growth factor concentrate are administered for the treatment of hair loss, each dose comprising at least 5ml of the growth factor concentrate.
- a method of preparing an intra- dermally, sub-dermally or topically administrable growth factor concentrate derived from human platelets comprising the following steps:
- cellular debris are separated from the thawed suspension and the resultant suspension of growth factors is diluted with an isotonic medium before sterile-filtering.
- a method of preparing an intra-dermally, sub-dermally or topically administrable growth factor concentrate derived from human platelets comprising the following steps:
- cellular debris are separated from the thawed suspension and the resultant suspension of growth factors is diluted with an isotonic medium before sterile-filtering.
- a method of preparing an intra-dermally, sub-dermally or topically administrable growth factor concentrate derived from human platelets comprising the following steps:
- a fixed number of platelets is suspended in a fixed volume of the isotonic medium to obtain the required concentration of growth factors in the growth factor concentrate;
- the snap-freezing is carried out at a temperature of -120°C to -200°C;
- the thawing is carried out at 25°C to 37°C;
- cellular debris are separated from the thawed suspension and the resultant suspension of growth factors is diluted with the isotonic medium before sterile-filtering and lyophilized with excipients after sterile-filtering,
- the isotonic medium in step (a) is plasma
- the volume of plasma does not exceed 5 ml.
- Freshly collected platelets or fresh blood can be collected from donors or even blood banks for large scale manufacturing of GFC.
- the blood is preferably transported to the central processing laboratory at 20-24°C in a transportation box.
- Blood can alternatively be collected from donors requiring treatment with the GFC.
- Blood may also be collected from other mammalian species such as horse, dog, cat, buffalo, cow, sheep, goat, rodents etc. from either jugular vein or the cephalic vein or femoral vein.
- One part of the blood sample collected is preferably routinely* processed for complete blood count (CBC), and rapid infectious disease marker testing for Human Immunodeficiency Virus (HIV- 1,2), Hepatitis B virus (HBV), Hepatitis C Virus (HCV), Venereal disease research laboratory (VDRL) tests.
- the remaining part of the blood collected is preferably sent to a class B environment clean-room for further processing to yield the GFC.
- the clean room temperature is preferably maintained at 22°C with a relative humidity of 55 %.
- the blood sample to be used for obtaining the GFC is centrifuged at 109 g to 680 g, preferably at. about 382 g for 15 minutes at 22°C for isolation of platelets.
- a top layer of yellow coloured platelet rich plasma PRP
- WBC white blood corpuscles
- RBC red blood corpuscles
- the top layer is aspirated carefully to maximize platelet yield, while ensuring that no WBCs are picked up, and placed in another sterile centrifuge tube.
- the platelet rich plasma collected is then centrifuged at 680 to 3442 g, preferably at about 2720 g for 10 minutes. This separates the PRP into a platelet pellet and a platelet poor plasma (PPP) supernatant.
- PPP platelet poor plasma
- Platelets of any desired concentrations could be prepared in this manner which is not possible in other known devices for obtaining PRP.
- the platelet pellet is re-suspended in the appropriate amount of isotonic medium.
- the isotonic medium is platelet free plasma, platelet poor plasma, multiple electrolyte isotonic solution or combinations thereof.
- the concentrated platelet pellet can be re-suspended in 1 to 10 ml of multiple electrolyte isotonic solution or platelet free plasma which is tested for ABO compatibility before use.
- the platelets are then counted and further isotonic medium can be added so that the platelet numbers are adjusted to the count of approximately 250 to 5000 ⁇ 10 6 platelets per ml; preferably approximately 875 ⁇ 10 6 platelets per ml.
- This platelet suspension is then subjected to a physiological activation by freezing at -196°C.
- the centrifuge tube containing the concentrated platelet suspension is placed in liquid nitrogen for 120 seconds and then subjected to a rapid thawing. Rapid thawing is done at 37°C for 120 seconds.
- Just one freeze-thaw cycle is sufficient to physiologically activate the platelets and causes lysis of the platelet membranes. This freeze- thaw cryo-stimulates the platelets to release growth factors.
- the suspension can then be mixed with 4 to 14 ml of PPP removed at the earlier step or platelet free plasma or multiple electrolyte isotonic solution and subjected to high speed centrifugation at a speed of 17610 g for 30 minutes.
- the last centrifugation is critical to remove all the plasma membranes or membrane antigens of platelets or debris so as to obtain an acellular solution.
- high speed centrifugation supernatant is aspirated and transferred to another sterile tube.
- the GFC is amenable to lyophilisation after mixing the GFC with 2 to 10% mannitol, sucrose and/or glycine added to the GFC as bulking agent/lyoprotectant for lyophilisation.
- the lyophilised product is sealed with a flip off cap for clinical application and is stable for more than one year at 4°C.
- Lyophilised GFC can be reconstituted with 5-15 ml of multiple electrolyte isotonic solution.
- Sterile water for injection can also be used as a diluent for reconstitution, with 1% human serum albumin.
- Blood type matched plasma can also be used as a diluent.
- the platelet pellet is suspended in 1ml of multiple electrolyte isotonic solution and after the freeze-thaw, the thawed solution is then mixed with 9 ml of plasma which had been removed from the upper layer at the end of the second centrifugation.
- the multiple electrolyte isotonic solution is supplemented with pharmaceutically acceptable excipients.
- the pharmaceutically acceptable additives are selected from a group comprising of Acid Citrate Dextrose-solution A (ACD-A), Ethylenediaminetetraacetic acid (EDTA), and Citrate phosphate dextrose adenine (CPDA).
- GFC comprises a combination of Growth Factors such as EGF, VEGF, TGF- ⁇ , bFGF, Insulin like growth factor- 1 (IGF-1), Hepatocyte Growth Factor (HGF), Platelet derived growth Factor- (PDGF-AA), (PDGF-AB), (PDGF-BB); Cytokines like RANTES, Interleukin-1 beta (IL- ⁇ ⁇ ), Macrophages inhibitory protein-1 alpha ( ⁇ - ⁇ ⁇ ), GRO-alpha, ENA-78, MCP-3, NCP, IGFBP-3; Basic Proteins like Platelet factor 4 (PF-4), Endostatin, PBP, Connective tissue activating peptide(CTAP), Neutrophil activating peptide (NAP); adhesive proteins such as ECGF, Plasminogen activator inhibitior-1 , Laminin-8, Fibrinogen, Fibronectin, Thrombospondin and antimicrobial agents like Thrombocidin.
- IGF-1 growth factor- 1
- HGF Hepat
- the GFC product was tested for endotoxin so as to confirm that the product was safe and free from bacterial contamination.
- the GFC product was also tested for infectious diseases like HIV, HCV, HBV, Syphilis etc and was found to be disease-free and safe for clinical applications.
- the snap-freezing is done in liquid nitrogen or in liquid helium.
- the thawing can be done in a sterile water bath at 37°C.
- the isotonic medium can be multiple electrolyte isotonic solution, plasma, platelet free plasma, platelet poor plasma or a combination thereof.
- the multiple electrolyte isotonic solution can be supplemented with pharmaceutically acceptable excipients.
- the excipients include mannitol, sucrose, glycine or combinations thereof.
- the cellular debris are removed from the thawed suspension by centrifuging the thawed suspension at 1 1270 g to 17610 g for 25 to 35 minutes and isolating the supernatant.
- the platelets of step (a) of the method can be obtained by plateletpheresis, or by centrifugation of whole blood.
- the platelets of step (a) are preferably obtained by:
- centrifuging at least 10 ml of anticoagulated human blood at 109 g to 680 g for 5 to 20 minutes;
- step (b) isolating the plasma-free pellet of platelets obtained at the end of step (b). Still preferably, the centrifugation in step (a) is carried out at 382 g for 15 minutes and the centrifugation in step (b) is carried out at 2720 g for 10 minutes.
- a method of treating dermatological conditions comprising administering topically, intra-dermally or sub-dermally the growth factor concentrate prepared by any of the methods above.
- the growth factor concentrate prepared by any of the methods above for treatment of human dermatological, conditions.
- the conditions can include androgenetic alopecia and hair loss.
- androgenetic alopecia and hair loss are treated with the growth factor concentrate derived from approximately 875 x 10 6 human platelets per ml.
- a therapeutic composition for topical, sub-dermal or intra-dermal administration comprising the growth factor concentrate prepared by any of the methods above in combination with supplemental constituents including blood, saline, silver nanoparticles, hyaluronic acid, immuno-modulatory peptides, growth factors, hormones, antibiotics, monoclonal antibodies, recombinant receptors, carriers or combinations thereof.
- the composition can be in the form of a cream, gel, aqueous solution, spray-aerosol or transdermal patch.
- the composition has about 500 to 1500 million platelets per ml preferably 875 million platelets per ml as a therapeutic dose for treatment of hair loss.
- Example 1 In order that those skilled in the art will be better able to practice the present disclosure, the following examples are given by way of illustration and not by way of limitation.
- vacutainers Human blood was withdrawn into vacutainers after getting informed consent from the patient. About 50-60 ml blood is collected in two types of vacutainers: 50 ml in vacutainers containing ACD-A for preparing GFC and 5-10 ml blood in EDTA tubes for infectious disease marker testing and complete blood parameter testing. Minimum 50 ml of blood was taken for 15 ml of GFC preparation. Blood was transported at 15 to 30°C preferably at 22°C within 4 hours of withdrawal. The first centrifugation was done at 382 g for 15 minutes since platelet recovery at 382 g for 15 minutes is optimum with a loss of only 8-10% platelets Platelet loss is significantly greater at lower or higher centrifugation speeds.
- the solution was then snap-frozen irliliquid nitrogen at -196°C for 2 minutes.
- the frozen solution was then rapidly thawed in a water bath at 37°C for 2 minutes.
- the thawed solution was then mixed with 14 ml of PPP. This solution was then transferred to another sterile tube and subjected to high speed centrifugation at 17610 g for 30 minutes.
- the supernatant containing the growth factors was collected and sterile filtered through a 0.22micron filter.
- Inclusion Criteria Male subjects were recruited in the study. Inclusion and exclusion criteria were as follows: Inclusion Criteria:
- Subjects with alopecia areata, alopecia totalis, alopecia universalis, alopecia diffusa and cicatricial alopecia
- Subjects with dermatological disorder of scalp that might interfere with study evaluation 4.
- Evidence of hypogonadism defined as serum testosterone ⁇ 250 ng/dl 5.
- the GFC of the present invention is derived from predetermined numbers of platelets yielding proportionate amounts of growth factors which in turn serves to provide consistent clinical results.
- Fixed dosages of GFC can be administered to patients to treat AGA or hair loss. Further, less than 10% platelets are lost by the method of the invention.
- the platelets are physiologically activated without incorporating any additional materials or chemical substances, like calcium chloride or bovine thrombin, and hence it is safe.
- the single freeze-thaw makes the process less time consuming and more suitable for large scale production and the growth factors are not denatured as in the case of multiple freeze-thaws.
- the present invention provides GFCs that can have standardized concentrations which can be diluted as per the requirement to make it suitable for specific clinical indications. Another benefit of the present method is that multiple doses of GFC can be prepared from a single blood draw and hence this method is cost effective and not time consuming.
- GFC does not show any flocculation on long-term storage for upto six months at -20°C; A small floe is sometimes seen which generally gets dissolved within 2- 3 minutes at room temperature and hence it is possible to use GFC as an "off the shelf product, which can be stored for upto six months without any problem of losing its potency. Also, since the GFC is acellular and devoid of plasma membranes or other antigenic materials, it does not elicit any immune reactions or formation of allo-antibodies. GFC can optionally be made plasma free so that it can be used as a therapeutic agent without any problem of ABO incompatible plasma that may cause immune reactions.
- GFC is also amenable to lyophilisation so that the GFC can be stored at room temperature or in a 4°C refrigerator without any degradation for more than one year. Further, lyophilized GFC can be made into a cream, gel, aqueous solution, spray-aerosol or transdermal patch. All in all, GFC is a natural i.e. non-recombinant product and the method provided by the present invention for production of GFC is economical. Further, GFC shows improved clinical outcomes due to the significantly higher level of growth factors in GFC as compared to multiple freeze-thawed HPL prepared by known methods. GFC also serves as a personalised therapy for patients requiring specific concentrations of GFC to be administered as the GFC can be prepared in any desired concentration.
- GFC contains therapeutic level of growth factors that is specifically suitable for hair loss. Injection of GFC at the affected part improves the hairline and density and the caliber of total hair.
- GFC derived from the platelets contains several important growth factors, cytokines, chemokine, anti-microbial agents and basic proteins and has very good therapeutic potential for various indications including AGA.
- the dosage of GFC administered and method of administration according to an embodiment of the present invention result in significant reduction in hair loss and also stimulate growth of hair.
- Growth factors present in GFC such as PDGF-AB, VEGF, TGF- ⁇ , EGF, bFGF, help in reducing inflammation, strengthening vasculature, stimulate extracellular signal regulated kinase (ERK) and AKT signalling.
- ERK and AKT signalling increases level of growth factor like FGF-7 and beta-catenin which is a potent stimulator for hair growth.
- growth factors proliferate dermal papillae (DP) and promotes faster transition of telogen to anagen phase.
- VEGF helps in strengthening vasculature which helps to provide nutrients to follicles.
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Abstract
The invention relates to an intra-dermally, sub-dermally or topically adrainistrable growth factor concentrate derived from approximately 500 x l06 to 1500 χ 106 human platelets per ml for treating hair loss. The concentrate comprises approximately 800 to 1200 pg/ml of Epidermal growth factor (EGF), 20 to 80 pg/ml of Vascular Endothelial growth factor (VEGF), 15 to 30 pg/ml of Basic fibroblast growth factor (b-FGF), 30000 to 40000 pg/ml of Transforming growth factor-β (TGF- β) and 100000 to 200000 pg/ml of Platelet Derived growth factor- AB (PDGF- AB).
Description
TITLE OF THE INVENTION A growth factor concentrate for treating hair loss FIELD OF THE INVENTION
This invention relates to a growth factor concentrate for treating hair loss.
BACKGROUND OF THE INVENTION
Hair loss due to androgenetic alopecia (AGA), or pattern alopecia, is a common disorder affecting both men and women. The incidence of androgenetic alopecia is generally considered to be greater in males than in females, although some evidence suggests that the apparent differences in incidence may be a reflection of different expression in males and females. This condition is also known as Male Pattern Hair Loss (MPHL) in men and Female Pattern Hair Loss (FPHL) in women. Alopecia in these cases is characterized by thinning of hair as opposed to follicular loss.
In men, MPHL does not appear until after puberty, usually manifesting in the third decade and in almost all cases by the fourth decade of life. MPHL typically begins with bi-temporal recession, followed by progressive thinning in the frontal and vertex areas of the scalp and recession of the frontal hairline is common. Over time, the frontal and vertex thinning areas may merge, resulting in near complete visible hair loss over the top of the scalp. In contrast, FPHL may present as late as the sixth decade of life and is characterized by diffuse thinning in the frontal and parietal areas of the scalp with preservation of the frontal hairline being the norm.
A variety of genetic and possibly even environmental factors appear to play a role in AGA. Although researchers have long studied the factors that may contribute to AGA, these remain largely unknown. Minimally, pattern hair loss is related to hormones called androgens, particularly dihydrotestosterone (DHT). Androgens are important for normal male sexual
development before birth and during puberty. Androgens also have other important functions in both males and females, such as regulating hair growth and sex drive.
MPHL is caused by a genetic sensitivity of hair follicles to DHT. This hormone causes follicles to shrink or "miniaturize". In turn, this shortens their lifespan and prevents them from producing hair normally. Recently the existing theories have been challenged on the ground that while the androgens in question are responsible for hair growth on the face and all over the body of men, hair loss only occurs at the top of the scalp. For example, it has been suggested that AGA is a consequence of the anabolic effect of androgens such as hormonal changes leading to structural changes in skin and scalp which in turn cause hair loss.
Current available treatment for AGA is topical application of minoxidil. It is available for the treatment of AGA in both men and women. The other treatment option available is oral finasteride which is less extensively evaluated. Extensive clinical trials on men with AGA have shown that minoxidil treatment is only effective in less than half the cases. For example, the conclusion of a year-long, randomised, double-blind trial involving 56 males, was that cosmetically acceptable hair growth occurred in 32% of individuals (De Villez RL. Topical minoxidil therapy in hereditary androgenetic alopecia. Arch Dermatol 1985; 121 : 197-202). In another 12-month study, minoxidil produced at least moderate cosmetic improvement in only 24% of individuals (Olsen E, Weiner MS, DeLong ER, et al. Topical minoxidil in early male pattern baldness. J Am Acad Dermatol 1985; 13: 185-92). Data derived from multiple, double- blind, placebo controlled trials, involving over 2000 males between the ages of 18 and 50 years, showed that twice daily application of 2% minoxidil solution over one year produced moderate to dense re-growth in approximately 30 to 35% of patients (Olsen E. Androgenetic alopecia. In: Olsen E, editor. Disorders of hair growth. New York: McGraw-Hill Inc., 1994: 257). Since
minoxidil is also a vasodilator, it is speculated that by widening blood vessels and opening potassium channels, it allows more oxygen, blood and nutrients to reach the follicle. This effect of minoxidil can also cause follicles in the telogen phase to shed, usually soon to be replaced by new, thicker hair (in a new anagen phase). However, minoxidil has severe side effects. The alcohol present in topical preparations may dry the scalp, resulting in dandruff. Side effects of oral minoxidil may include swelling of the face and extremities, rapid and irregular heartbeat, light headedness, cardiac lesions, and focal necrosis of the papillary muscle and subendocardial areas of the left ventricle. There have been cases of allergic reactions to minoxidil or the non- active ingredient propylene glycol, which is found in some forms of topical minoxidil. Pseudoacromegaly is also a rarely reported side effect of minoxidil.
In late 1997, the US Food and Drug Administration (US FDA) approved oral finasteride at a dosage of 1 mg/day for the treatment of AGA in men (Davies K. Hair apparent Nature 1998; 391 : 537-9). The drug is not licensed for administration to women because of the potentially teratogenic effects of finasteride on a male fetus, should the woman be pregnant. Oral finasteride showed improvement in 48% treated males as against 7% who were given a placebo. The results showed that finasteride treatment over a 1-year period resulted in a significant hypertrichotic response at the affected regions. Finasteride is a azasteroid and since the drug does not cure the underlying genetic causes of AGA, ceasing daily administration, results in the shedding of regrown hair and the resumption of hair loss. Therefore, continual use of finasteride is necessary . if the cosmetic benefit is to be sustained. Finasteride is usually well tolerated. However, there is a small risk of transient impotence as an adverse effect, which is reversible upon treatment cessation (Amichai B, Grunwald MH, Sobel R. 5a-reductase inhibitors: a new hope in dermatology? Int J Dermatol 1997; 36: 182-4).
Zimber et. al., studied the effect of a bioengineered, non-recombinant, human cell derived formulation termed hair stimulating complex (HSC) containing growth factors to assess its effect on hair growth activity in MPHL. HSC showed in vitro Wnt activity and contained Follistatin, Keratinocyte Growth Factor (KGF) and Vascular Endothelial Growth Factor (VEGF). In the placebo controlled randomized single site trial, a single intradermal injection of HSC in 26 patients showed significant improvement in hair growth over the placebo on 12 and 52 weeks. There was increase in hair shaft thickness by 6.3%, thickness density at 12.8% and terminal hair density at 20.6%. After one year, total cell count continued to increase. Results concluded that a single intra-dermal injection is proved to be beneficial for hair growth (Zimber MP, Ziering C, Zeigler F, Hubka M, Mansbridge JN, Baumgartner M, Hubka K, Keliar R, Perez-Meza D, Sadick N, Naughton GK. (201 1 ) Hair regrowth following a Wnt- and Follistatin containing treatment: safety and efficacy in a first in man Phase 1 clinical trial. J Drug Dermatol, 10: 1308-1312).
SUMMARY OF THE INVENTION
According to an embodiment of the invention there is provided an intra-dermally, sub-dermally or topically administrable growth factor concentrate derived from approximately 500 l O6 to 1500 x 106 human platelets per ml for treating hair loss, the concentrate comprising approximately 800 to 1200 pg/ml of Epidermal growth factor (EGF), 20 to 80 pg/ml of Vascular Endothelial growth factor (VEGF), 15 to 30 pg/ml of Basic fibroblast growth factor (b-FGF), 30000 to 40000 pg/ml of Transforming growth factor-β (TGF- β) and 100000 to 200000 pg/ml of Platelet Derived growth factor-AB (PDGF-AB).
According to another embodiment of the invention there is provided a lyophilized, intra-dermally, sub-dermally or topically administrable growth factor concentrate derived from approximately 500 x l O6 to 1500 x l O6 human platelets per ml for treating hair loss, the concentrate comprising
800 to 1200 pg/ml of EGF, 20 to 80 pg/ml of VEGF, 15 to 30 pg/ml of b-FGF, 30000 to 40000 pg/ml of TGF- β and 100000 to 200000 pg/ml of PDGF-AB suspended in an isotonic medium along with excipients for lyophilisation. According to yet another embodiment of the invention there is provided a therapeutic composition for intra-dermal, sub-dermal or topical administration comprising the growth factor concentrate in combination with supplemental constituents including blood, saline, silver nanoparticles, hyaluronic acid, immuno-modulatory peptides, growth factors, hormones, antibiotics, monoclonal antibodies, recombinant receptors, carriers or combinations thereof.
'
According to still another embodiment of the invention there is provided a method of treatment of hair loss comprising injecting a growth factor concentrate into the scalp first into the subcutaneous fat followed by deep dermal injection and lastly sub-dermal injection, wherein the growth factor concentrate is derived from approximately 875 * 106 human platelets per ml for treating hair loss, the concentrate comprising 800 to 1200 pg/ml of Epidermal growth factor (EGF), 20 to 80 pg/ml of VEGF, 15 to 30 pg/ml of b-FGF, 30000 to 40000 pg/ml of TGF- β and 100000 to 200000 pg/ml of PDGF-AB. The growth factor concentrate may also be administered by micro-needling or by meso-gun or a combination of the above methods of administration. It is to be understood that both the foregoing general description and the following detailed description of the present embodiments of the invention are intended to provide an overview or framework for understanding the nature and character of the invention as it is claimed. The accompanying graphical representations are included to substantiate the invention and are incorporated into and constitute a part of this specification.
BRIEF DESCRIPTION OF THE DRAWINGS
In order that the disclosure may be readily understood and put into practical effect, reference will now be made to exemplary embodiments as illustrated with reference to the accompanying figures. The figures together with a detailed description below, are incorporated in and form part of the specification, and serve to further illustrate the embodiments and explain various principles and advantages, in accordance with the present disclosure where:
FIG. lA and FIG IB respectively show a photographic comparison between hair growth before treatment and at two months after administration of the 3rd dose of growth factor concentrate (GFC), the treatment consisting of three intra-dermal injections of GFC derived from 875 * 106 platelets per ml, each subsequent injection being given at an interval of one month from the previous injection.
FIG. 2 shows total hair count per sq. cm. of ten patients' scalps using a folliscope before treatment and at two months and at four months after administration of the 3rd dose of GFC, each subsequent injection being given at an interval of one month from the previous injection.
FIG. 3 shows the caliber of ten patients' hair before treatment and at two months and at four months after administration of the 3rd dose of GFC, each subsequent injection being given at an interval of one month from the previous injection.
FIG. 4 shows the mean total hair count per sq. cm. of ten patients' scalps using a folliscope before treatment and at two and four months after administration of the 3rd dose of GFC, each subsequent injection being given at an interval of one month from the previous injection.
FIG. 5 shows the mean caliber of ten patients' hair before treatment and at two and four months after administration of the 3rd dose of GFC, each subsequent injection being given at an interval of one month from the previous injection. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
For simplicity and illustrative purposes, the present invention is described by referring mainly to exemplary embodiments thereof. In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention. It will be apparent, however, to one of ordinary skill in the art that the present invention may be practiced without limitation to these specific details. In other instances, well known methods have not been described in detail so as not to unnecessarily obscure the present invention.
In the context of the invention, the term "growth factor concentrate" or "GFC" as used in the specification refers to a standardized concentration of growth factors prepared according to an embodiment of the invention, the growtli factors being derived from the cryo-stimulation of platelets which have been counted, the platelets being sourced from human blood. The GFC also contains cytokines, chemokines, adhesive proteins and other modulatory peptides.
Also, "multiple electrolyte isotonic solution" as used in the specification in the context of the invention comprises sodium chloride, (NaCl); sodium gluconate (C6HnNa07); sodium acetate trihydrate, (C2H3Na02 »3H20); potassium chloride, (KC1); and magnesium chloride, (MgCl2'6H20) and human serum albumin. Preferably, each 100 ml of multiple electrolyte isotonic solution contains 526 mg of sodium chloride, (NaCl); 502 mg of sodium gluconate (C6HnNa07); 368 mg of sodium acetate trihydrate, (C2H3Na02*3H20); 37 mg of potassium chloride, (KC1); and 30 mg of magnesium chloride, (Mg02 «6H20) and 1 to 5% of human serum
albumin. The pH is adjusted with sodium hydroxide to 7.4 (or 6.5 to 8.0), sterile filtered and tested for endotoxin.
Further, the term "platelet free plasma" as used in the specification in the context of the invention comprises the supernatant collected from plasma that has been centrifuged at about 17610 g and then sterile filtered. The term "platelet poor plasma" or "PPP" as used in the specification in the context of the invention comprises supernatant collected from whole blood that has been centrifuged at about 2720 g. Also, the term "baseline" as used in the specification in the context of the invention refers to the hair count of a study subject prior to administration of GFC according to an embodiment of the invention.
Preferably, the isotonic medium in the growth factor concentrate is multiple electrolyte isotonic solution, plasma, platelet free plasma, platelet poor plasma or a combination thereof.
Preferably, the concentrate is derived from approximately 875 x lO6 human'platelets per ml.
The growth factor concentrate according to an embodiment of the invention is used for treatment of hair loss, particularly for treatment of androgenetic alopecia.
The therapeutic composition according to an embodiment of the invention can be in the form of a cream, gel, aqueous solution, spray-aerosol or transdermal patch.
Preferably, three doses of the growth factor concentrate are administered for the treatment of hair loss, each dose comprising at least 5ml of the growth factor concentrate.
According to an embodiment of the invention there is provided a method of preparing an intra- dermally, sub-dermally or topically administrable growth factor concentrate derived from human platelets comprising the following steps:
a. suspending human platelets in multiple electrolyte isotonic solution;
b. snap-freezing the suspension;
c. thawing the frozen suspension; and
d. sterile-filtering the suspension
wherein, a fixed number of platelets is suspended in a fixed volume of multiple electrolyte isotonic solution to obtain the required concentration of growth factors in the growth factor concentrate,
snap-freezing of the suspension is carried out at a temperature of -120°C to -200°C, thawing of the frozen suspension is carried out at 25°C to 37°C, and
cellular debris are separated from the thawed suspension and the resultant suspension of growth factors is diluted with an isotonic medium before sterile-filtering.
According to yet another embodiment of the invention there is provided a method of preparing an intra-dermally, sub-dermally or topically administrable growth factor concentrate derived from human platelets comprising the following steps:
a. suspending human platelets in upto 5ml of plasma;
b. snap-freezing the suspension;
c. thawing the frozen suspension; and
d. sterile-filtering the suspension
wherein, a fixed number of platelets is suspended in a fixed volume of plasma to obtain the required concentration of growth factors in the growth factor concentrate,
snap-freezing of the suspension is carried out at a temperature of -120°C to -200°C, thawing of the frozen suspension is carried out at 25°C to 37°C, and
cellular debris are separated from the thawed suspension and the resultant suspension of growth factors is diluted with an isotonic medium before sterile-filtering.
According to a further embodiment of the invention there is provided a method of preparing an intra-dermally, sub-dermally or topically administrable growth factor concentrate derived from human platelets comprising the following steps:
a. suspending human platelets in an isotonic medium;
b. snap-freezing the suspension;
c. thawing the frozen suspension; and
d. sterile-filtering the suspension
wherein, a fixed number of platelets is suspended in a fixed volume of the isotonic medium to obtain the required concentration of growth factors in the growth factor concentrate;
the snap-freezing, is carried out at a temperature of -120°C to -200°C;
the thawing is carried out at 25°C to 37°C; and
cellular debris are separated from the thawed suspension and the resultant suspension of growth factors is diluted with the isotonic medium before sterile-filtering and lyophilized with excipients after sterile-filtering,
provided that where the isotonic medium in step (a) is plasma, the volume of plasma does not exceed 5 ml.
According to yet another embodiment of the invention there is provided an intra-dermally, sub- dermally or topically administrable growth factor concentrate derived from a fixed number of human platelets, the concentrate comprising the required concentration of growth factors suspended in an isotonic medium.
According to still another embodiment of the invention there is provided a lyophilized, intra- dermally, sub-dermally or topically administrable growth factor concentrate derived from human platelets, the concentrate comprising the required concentration of growth factors suspended in an isotonic medium along with excipients for iyophilisation.
According to yet another embodiment of the invention there is provided a dosage of an intra- dermally, sub-dermally or topically administrable growth factor concentrate derived from approximately 875 x l O6 human platelets per ml, the concentrate comprising approximately 800 to 1200 pg/ml of EGF, 20 to 80 pg/ml of VEGF, 15 to 30 pg/ml of b-FGF, 30000 to 40000 pg/ml of TGF- β and 100000 to 200000 pg/ml of PDGF-AB.
Freshly collected platelets or fresh blood can be collected from donors or even blood banks for large scale manufacturing of GFC. The blood is preferably transported to the central processing laboratory at 20-24°C in a transportation box. Blood can alternatively be collected from donors requiring treatment with the GFC. Blood may also be collected from other mammalian species such as horse, dog, cat, buffalo, cow, sheep, goat, rodents etc. from either jugular vein or the cephalic vein or femoral vein. One part of the blood sample collected is preferably routinely* processed for complete blood count (CBC), and rapid infectious disease marker testing for Human Immunodeficiency Virus (HIV- 1,2), Hepatitis B virus (HBV), Hepatitis C Virus (HCV), Venereal disease research laboratory (VDRL) tests. The remaining part of the blood collected is preferably
sent to a class B environment clean-room for further processing to yield the GFC. The clean room temperature is preferably maintained at 22°C with a relative humidity of 55 %.
Stability of the platelets in the whole blood (in terms of growth factor levels) was checked for different temperatures and time points and it was found that platelets in whole blood are stable between 15 to 30°C until 24 hours for the purpose of GFC preparation i.e. the growth factor levels measured using ELISA after recovering platelets at different temperatures and time points remained stable upto 24 hours. According to an embodiment of the invention, the blood sample to be used for obtaining the GFC is centrifuged at 109 g to 680 g, preferably at. about 382 g for 15 minutes at 22°C for isolation of platelets. After centrifugation, three layers are observed: a top layer of yellow coloured platelet rich plasma (PRP), a middle layer of white blood corpuscles (WBC) and the bottom layer of red blood corpuscles (RBC). The top layer is aspirated carefully to maximize platelet yield, while ensuring that no WBCs are picked up, and placed in another sterile centrifuge tube. The platelet rich plasma collected is then centrifuged at 680 to 3442 g, preferably at about 2720 g for 10 minutes. This separates the PRP into a platelet pellet and a platelet poor plasma (PPP) supernatant. The entire PPP is collected in a sterile centrifuge tube and stored at room temperature for later use. Platelets of any desired concentrations could be prepared in this manner which is not possible in other known devices for obtaining PRP. For autologous use, the platelet pellet is re-suspended in the appropriate amount of isotonic medium. Preferably, the isotonic medium is platelet free plasma, platelet poor plasma, multiple electrolyte isotonic solution or combinations thereof. For allogeneic purposes, the concentrated platelet pellet can be re-suspended in 1 to 10 ml of multiple electrolyte isotonic solution or platelet free plasma which is tested for ABO compatibility before use. The platelets are then counted and further isotonic medium can be added so that the platelet
numbers are adjusted to the count of approximately 250 to 5000 χ 106 platelets per ml; preferably approximately 875 χ 106 platelets per ml. This platelet suspension is then subjected to a physiological activation by freezing at -196°C. The centrifuge tube containing the concentrated platelet suspension is placed in liquid nitrogen for 120 seconds and then subjected to a rapid thawing. Rapid thawing is done at 37°C for 120 seconds. Just one freeze-thaw cycle is sufficient to physiologically activate the platelets and causes lysis of the platelet membranes. This freeze- thaw cryo-stimulates the platelets to release growth factors. The suspension can then be mixed with 4 to 14 ml of PPP removed at the earlier step or platelet free plasma or multiple electrolyte isotonic solution and subjected to high speed centrifugation at a speed of 17610 g for 30 minutes. The last centrifugation is critical to remove all the plasma membranes or membrane antigens of platelets or debris so as to obtain an acellular solution. After high speed centrifugation supernatant is aspirated and transferred to another sterile tube. The GFC is amenable to lyophilisation after mixing the GFC with 2 to 10% mannitol, sucrose and/or glycine added to the GFC as bulking agent/lyoprotectant for lyophilisation. The lyophilised product is sealed with a flip off cap for clinical application and is stable for more than one year at 4°C. Lyophilised GFC can be reconstituted with 5-15 ml of multiple electrolyte isotonic solution. Sterile water for injection can also be used as a diluent for reconstitution, with 1% human serum albumin. Blood type matched plasma can also be used as a diluent. In an embodiment of the invention, the platelet pellet is suspended in 1ml of multiple electrolyte isotonic solution and after the freeze-thaw, the thawed solution is then mixed with 9 ml of plasma which had been removed from the upper layer at the end of the second centrifugation.
In a preferred embodiment of the invention, the multiple electrolyte isotonic solution is supplemented with pharmaceutically acceptable excipients. Preferably, the pharmaceutically
acceptable additives are selected from a group comprising of Acid Citrate Dextrose-solution A (ACD-A), Ethylenediaminetetraacetic acid (EDTA), and Citrate phosphate dextrose adenine (CPDA). GFC comprises a combination of Growth Factors such as EGF, VEGF, TGF-β, bFGF, Insulin like growth factor- 1 (IGF-1), Hepatocyte Growth Factor (HGF), Platelet derived growth Factor- (PDGF-AA), (PDGF-AB), (PDGF-BB); Cytokines like RANTES, Interleukin-1 beta (IL-Ι β), Macrophages inhibitory protein-1 alpha (ΜΙΡ-Ι α), GRO-alpha, ENA-78, MCP-3, NCP, IGFBP-3; Basic Proteins like Platelet factor 4 (PF-4), Endostatin, PBP, Connective tissue activating peptide(CTAP), Neutrophil activating peptide (NAP); adhesive proteins such as ECGF, Plasminogen activator inhibitior-1 , Laminin-8, Fibrinogen, Fibronectin, Thrombospondin and antimicrobial agents like Thrombocidin. These growth factors and other proteins have regenerative properties and help in initiating the healing process. The GFC product was tested for endotoxin so as to confirm that the product was safe and free from bacterial contamination. The GFC product was also tested for infectious diseases like HIV, HCV, HBV, Syphilis etc and was found to be disease-free and safe for clinical applications.
Clinical studies on hair loss were undertaken for the GFC prepared by the method of the present invention. In the clinical trials being conducted, administration of GFC led to a significant improvement in hair regeneration, and hair calibre.
It is within the scope of the invention to use other human cell types i.e. other than platelets as well, to yield the GFC according to the method of the invention. It is also within the scope of the
invention to use other mammalian cells or placenta to yield the GFC according to the method of the invention.
Preferably, the snap-freezing is done in liquid nitrogen or in liquid helium. The thawing can be done in a sterile water bath at 37°C. The isotonic medium can be multiple electrolyte isotonic solution, plasma, platelet free plasma, platelet poor plasma or a combination thereof. The multiple electrolyte isotonic solution can be supplemented with pharmaceutically acceptable excipients. Preferably, the excipients include mannitol, sucrose, glycine or combinations thereof. Preferably, the cellular debris are removed from the thawed suspension by centrifuging the thawed suspension at 1 1270 g to 17610 g for 25 to 35 minutes and isolating the supernatant.
The platelets of step (a) of the method can be obtained by plateletpheresis, or by centrifugation of whole blood. The platelets of step (a) are preferably obtained by:
a. centrifuging at least 10 ml of anticoagulated human blood at 109 g to 680 g for 5 to 20 minutes;
b. isolating the top-most layer containing platelets and centrifuging the same at 680 g to 3442 g for 5 to 15 minutes; and
c. isolating the plasma-free pellet of platelets obtained at the end of step (b). Still preferably, the centrifugation in step (a) is carried out at 382 g for 15 minutes and the centrifugation in step (b) is carried out at 2720 g for 10 minutes.
According to an embodiment of the invention, there is provided a method of treating dermatological conditions comprising administering topically, intra-dermally or sub-dermally the growth factor concentrate prepared by any of the methods above.
According to another embodiment of the invention, there is provided use of the growth factor concentrate prepared by any of the methods above for treatment of human dermatological, conditions. The conditions can include androgenetic alopecia and hair loss.
Preferably, androgenetic alopecia and hair loss are treated with the growth factor concentrate derived from approximately 875 x 106 human platelets per ml.
According to another embodiment of the invention, there is provided a therapeutic composition for topical, sub-dermal or intra-dermal administration comprising the growth factor concentrate prepared by any of the methods above in combination with supplemental constituents including blood, saline, silver nanoparticles, hyaluronic acid, immuno-modulatory peptides, growth factors, hormones, antibiotics, monoclonal antibodies, recombinant receptors, carriers or combinations thereof. The composition can be in the form of a cream, gel, aqueous solution, spray-aerosol or transdermal patch.
In an embodiment of the present disclosure, the composition has about 500 to 1500 million platelets per ml preferably 875 million platelets per ml as a therapeutic dose for treatment of hair loss.
In order that those skilled in the art will be better able to practice the present disclosure, the following examples are given by way of illustration and not by way of limitation.
Example 1
Human blood was withdrawn into vacutainers after getting informed consent from the patient. About 50-60 ml blood is collected in two types of vacutainers: 50 ml in vacutainers containing ACD-A for preparing GFC and 5-10 ml blood in EDTA tubes for infectious disease marker testing and complete blood parameter testing. Minimum 50 ml of blood was taken for 15 ml of GFC preparation. Blood was transported at 15 to 30°C preferably at 22°C within 4 hours of withdrawal. The first centrifugation was done at 382 g for 15 minutes since platelet recovery at 382 g for 15 minutes is optimum with a loss of only 8-10% platelets Platelet loss is significantly greater at lower or higher centrifugation speeds. Upon completion of first centrifugation three layers were formed. At the bottom were packed red blood cells, in the middle were leukocytes and the upper layer had plasma containing platelets. Plasma containing platelets was aspirated and transferred in another sterile tube. The second centrifugation was done at 2720 g for 10 minutes since centrifugation at 2720 g for 10 minutes results in platelet recovery of almost 99.5%. The upper layer had plasma and at the bottom was a packed platelet pellet. The entire plasma (PPP) was removed and placed in a sterile tube and stored at room temperature for later use. Platelet pellet was suspended in 1 ml of PPP. Platelets were then counted and the concentration of platelets per ml of plasma was noted to be 13125x 106. The solution was then snap-frozen irliliquid nitrogen at -196°C for 2 minutes. The frozen solution was then rapidly thawed in a water bath at 37°C for 2 minutes. In order to standardize the concentration of growth factors in the solution, the thawed solution was then mixed with 14 ml of PPP. This solution was then transferred to another sterile tube and subjected to high speed centrifugation at 17610 g for 30 minutes. The supernatant containing the growth factors was collected and sterile filtered through a 0.22micron filter.
Since the final volume of the GFC is 15 ml, and the number of platelets used for preparing the GFC were 13125 x 106, the effective concentration of platelets used for preparation of the GFC is 875 x l O6 platelets per ml. Example 2
Prospective, multi-centric, open label, randomized, bio-interventional, phase I/II pilot study to evaluate the safety and efficacy of the GFC prepared in Example 1 for treatment of AGA was carried out. The objective of this pilot study was to demonstrate the potential of autologous GFC prepared in Example 1 in the treatment of subjects with AGA. The primary objective was to assess the safety of autologous GFC in treatment of subjects with AGA. The secondary objective was to assess the efficacy of autologous GFC in treatment of subjects with AGA.
Male subjects were recruited in the study. Inclusion and exclusion criteria were as follows: Inclusion Criteria:
1 . Male subjects, aged between 18 to 50 years (both inclusive) and in general good health
2. Norwood- Hamilton grades III and IV AGA
3. Subjects willing to refrain from other AGA treatments during the entire study duration
4. Subjects who are willing to give informed consent and adhere to the study protocol.
Exclusion Criteria:
1. Subjects aged <18 or > 50 years
2. Subjects with alopecia areata, alopecia totalis, alopecia universalis, alopecia diffusa and cicatricial alopecia
3. Subjects with dermatological disorder of scalp that might interfere with study evaluation 4. Evidence of hypogonadism defined as serum testosterone < 250 ng/dl
5. Subjects with hair loss due to a certain medical aliment (such as typhoid) or due to some surgical intervention that might significantly affect the study outcome
6. Treatment with systemic or locally acting medication such as minoxidil treatment in the 6 month prior to study, finasterides 12 month prior to study
7. Subjects taking blood thinning medicines like Aspirin
8. Subjects on Anti-coagulant therapy
9. Subjects with clinically significant medical or psychiatric disease as determined by the investigator.
10. Subjects with history of malignancies
1 1. Subjects with immunocompromised systems
12. Subjects with autoimmune diseases
13. Subjects who are heavy smokers
14. Subjects with current or past (last 12 months) abuse of alcohol or drugs
15. Subjects unwilling to or unable to comply with the study protocol.
16. Subjects taking concomitant therapy that might interfere with the study results in investigators opinion or participating in another trial in the past 30 days.
After informed consent was taken, the patients were checked for inclusion and exclusion criteria before recruiting for the clinical study and were screened for Infectious Disease markers. A grid was made on the bald area and equidistant injection sites were marked out. Total 15 ml of GFC prepared as per Example 1 was administered in three doses of 5 ml each. Each dose was given intra-dermally in the scalp at the pre-designated injection sites. The area was disinfected thoroughly and pre-treated with a topical anaesthetic cream prior to injection. 1 ml of GFC was drawn into a 1 ml syringe with a 26 G needle. The needle was injected intra-dermally in the scalp deep into the dermis at a 45° angle. While retracting the needle 0.3ml to 0.4 ml was injected both
into the dermis and epidermis at each point. This procedure was repeated at different marked points. 5ml of GFC was administered at each visit. Totally 3 dosages were given at an interval of one month each. Folliscope analysis was done at baseline and at two and four months after the third injection. The total hair count per cm and the hair caliber was analyzed for ten study subjects at two months and at four months as shown in FIG. 2 and FIG. 3. Results shown in FIG. 2 and FIG. 3 show that the total hair count and hair caliber continued to improve even at four months, post treatment. Results shown in FIG. 4 and FIG. 5 demonstrate that there is a significant increase of total hair count as well as a marked improvement in hair caliber from the baseline. Statistical analysis was carried out by Repeated Measures ANOVA followed by the Dunnett's test.
As is evident from the above results, the GFC of the present invention is derived from predetermined numbers of platelets yielding proportionate amounts of growth factors which in turn serves to provide consistent clinical results. Fixed dosages of GFC can be administered to patients to treat AGA or hair loss. Further, less than 10% platelets are lost by the method of the invention. Also, the platelets are physiologically activated without incorporating any additional materials or chemical substances, like calcium chloride or bovine thrombin, and hence it is safe. Also, the single freeze-thaw makes the process less time consuming and more suitable for large scale production and the growth factors are not denatured as in the case of multiple freeze-thaws. Moreover, some clinical indications need a large amount of growth factors but some indications need very little growth factors since the presence of growth factor receptors vary from one cell type to another and one indication to another. Therefore, the present invention provides GFCs that can have standardized concentrations which can be diluted as per the requirement to make it suitable for specific clinical indications. Another benefit of the present method is that multiple doses of GFC can be prepared from a single blood draw and hence this method is cost effective and not time consuming. Further, GFC does not show any flocculation on long-term storage for
upto six months at -20°C; A small floe is sometimes seen which generally gets dissolved within 2- 3 minutes at room temperature and hence it is possible to use GFC as an "off the shelf product, which can be stored for upto six months without any problem of losing its potency. Also, since the GFC is acellular and devoid of plasma membranes or other antigenic materials, it does not elicit any immune reactions or formation of allo-antibodies. GFC can optionally be made plasma free so that it can be used as a therapeutic agent without any problem of ABO incompatible plasma that may cause immune reactions. GFC is also amenable to lyophilisation so that the GFC can be stored at room temperature or in a 4°C refrigerator without any degradation for more than one year. Further, lyophilized GFC can be made into a cream, gel, aqueous solution, spray-aerosol or transdermal patch. All in all, GFC is a natural i.e. non-recombinant product and the method provided by the present invention for production of GFC is economical. Further, GFC shows improved clinical outcomes due to the significantly higher level of growth factors in GFC as compared to multiple freeze-thawed HPL prepared by known methods. GFC also serves as a personalised therapy for patients requiring specific concentrations of GFC to be administered as the GFC can be prepared in any desired concentration.
GFC contains therapeutic level of growth factors that is specifically suitable for hair loss. Injection of GFC at the affected part improves the hairline and density and the caliber of total hair.. GFC derived from the platelets contains several important growth factors, cytokines, chemokine, anti-microbial agents and basic proteins and has very good therapeutic potential for various indications including AGA. As is evident, the dosage of GFC administered and method of administration according to an embodiment of the present invention result in significant reduction in hair loss and also stimulate growth of hair. Growth factors present in GFC such as PDGF-AB, VEGF, TGF-β, EGF, bFGF, help in reducing inflammation, strengthening vasculature, stimulate extracellular signal regulated kinase (ERK) and AKT signalling. Stimulation of ERK and AKT
signalling increases level of growth factor like FGF-7 and beta-catenin which is a potent stimulator for hair growth. These growth factors in turn proliferate dermal papillae (DP) and promotes faster transition of telogen to anagen phase. VEGF helps in strengthening vasculature which helps to provide nutrients to follicles.
What has been described and illustrated herein are preferred embodiments of the invention along with some of their variations. The terms, descriptions and figures used herein are set forth by way of illustration only and are not meant as limitations. Those skilled in the art will recognize that many variations are possible within the spirit and scope of the invention, which is intended to be defined by the claims in the complete specification— and their equivalents— in which all terms are meant in their broadest reasonable sense unless otherwise indicated.
Claims
1. An intra-dermally, sub-dermally or topically administrable growth factor concentrate derived from approximately 500 l O6 to 1500 x l O6 human platelets per ml for treating hair loss, the concentrate comprising approximately 800 to 1200 pg/ml of Epidermal growth factor (EGF), 20 to 80 pg/ml of Vascular Endothelial growth factor (VEGF), 15 to 30 pg/ml of Basic fibroblast growth factor (b-FGF), 30000 to 40000 pg/ml of Transforming growth factor-β (TGF- β) and 100000 to 200000 pg/ml of Platelet Derived growth factor-AB (PDGF-AB).
2. A lyophilized, intra-dermally, sub-dermally or topically administrable growth factor concentrate derived from approximately 500 x l O6 to 1500 x l O6 human platelets per ml for treating hair loss, the concentrate comprising 800 to 1200 pg/ml of EGF, 20 to 80 pg/ml of VEGF, 15 to 30 pg/ml of b-FGF, 30000 to 40000 pg/ml of TGF- β and 100000 to 200000 pg/ml of PDGF-AB suspended in an isotonic medium along with excipients for lyophilisation.
3. The concentrate as claimed in claim 2 wherein, the isotonic medium is multiple electrolyte isotonic solution, plasma, platelet free plasma, platelet poor plasma or a combination thereof.
4. The concentrate as claimed in claim 1 or 2 derived from approximately 875 x lO6 human platelets per ml.
5. Use of the growth factor concentrate as claimed in claim 1 or 2 for treatment of androgenetic alopecia and hair loss.
6. A therapeutic composition for intra-dermal, sub-dermal or topical administration for treating hair loss comprising the growth factor concentrate as claimed in claim 1 or 2 in combination with supplemental constituents including blood, saline, silver nanoparticles, hyaluronic acid, immuno-modulatory peptides, growth factors, hormones, antibiotics, monoclonal antibodies, recombinant receptors, carriers or combinations thereof.
7. The composition as claimed in claim 6 wherein, the composition is in the form of a cream, gel, aqueous solution, spray-aerosol or transdermal patch.
,
8. A method of treatment of hair loss comprising injecting a growth factor concentrate into the scalp first into the sub-cutaneous fat followed by deep dermal injection and lastly sub- dermal injection, wherein the growth factor concentrate is derived from approximately 875 x lO6 human platelets per ml for treating androgenetic alopecia, the concentrate comprising 800 to 1200 pg/ml of EGF, 20 to 80 pg/ml of VEGF, 15 to 30 pg/ml of b-FGF, 30000 to 40000 pg/ml of TGF- β and 100000 to 200000 pg/ml of PDGF-AB.
9. A method of treatment of hair loss comprising injecting a growth factor concentrate into the scalp by a meso-gun, wherein the growth factor concentrate is derived from approximately 875 x lO6 human platelets per ml for treating androgenetic alopecia, the concentrate comprising 800 to 1200 pg/ml of EGF, 20 to 80 pg/ml of VEGF, 15 to 30 pg/ml of b-FGF, 30000 to 40000 pg/ml of TGF- β and 100000 to 200000 pg/ml of PDGF- AB.
10. A method of treatment of hair loss comprising injecting a growth factor concentrate into the scalp by micro-needling, wherein the growth factor concentrate is derived from approximately 875 x lO6 human platelets per ml for treating androgenetic alopecia, the concentrate comprising 800 to 1200 pg/ml of EGF, 20 to 80 pg/ml of VEGF, 15 to 30 pg/ml of b-FGF, 30000 to 40000 pg/ml of TGF- β and 100000 to 200000 pg/ml of PDGF- AB.
1 1. The method as claimed in claim 8 or 9 or 10, wherein three doses of the growth factor concentrate are administered for the treatment of hair loss, each dose comprising at least 5ml of the growth factor concentrate.
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Cited By (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2016116560A1 (en) | 2015-01-21 | 2016-07-28 | Turkof Edvin | Platelet concentrate for increase of cell regeneration and cell growth |
WO2018015973A1 (en) | 2016-07-21 | 2018-01-25 | Khorakiwala Habil F | Topical pharmaceutical composition for promoting hair growth and/or reducing hair loss |
CN109310711A (en) * | 2016-03-23 | 2019-02-05 | 里尔大学中心医院 | The preparation of pellet platelets lysate and its purposes for the treatment of the nervous system disease |
AU2015388056B2 (en) * | 2015-03-23 | 2019-04-04 | HCell Inc. | Method for promoting hair growth and preventing hair loss of patient, composition therefor, and method for preparing composition |
US11083687B2 (en) * | 2016-06-03 | 2021-08-10 | Embryogenesis Pty Ltd | Skin care formulation |
WO2022035849A1 (en) * | 2020-08-10 | 2022-02-17 | Venturis Therapeutics, Inc. | Methods reducing hair loss and/or increasing hair regrowth |
CN116831983A (en) * | 2023-09-02 | 2023-10-03 | 潍坊市人民医院(潍坊市公共卫生临床中心) | Concentrated growth factor composite gel preparation, preparation method and hair growth application |
Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20060142198A1 (en) * | 2004-07-02 | 2006-06-29 | Wound Care Partners Llc | Compositions for treating wounds and processes for their preparation |
US20100030152A1 (en) * | 2007-05-31 | 2010-02-04 | Mcure Co., Ltd. | Multi Injection Microneedle Therapy System |
US20110294731A1 (en) * | 2009-02-13 | 2011-12-01 | Invitrx, Inc. | Skin cream |
-
2013
- 2013-04-22 WO PCT/IN2013/000268 patent/WO2014027363A1/en active Application Filing
Patent Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20060142198A1 (en) * | 2004-07-02 | 2006-06-29 | Wound Care Partners Llc | Compositions for treating wounds and processes for their preparation |
US20100030152A1 (en) * | 2007-05-31 | 2010-02-04 | Mcure Co., Ltd. | Multi Injection Microneedle Therapy System |
US20110294731A1 (en) * | 2009-02-13 | 2011-12-01 | Invitrx, Inc. | Skin cream |
Non-Patent Citations (1)
Title |
---|
BORZINI ET AL: "PLATELET-RICH PLASMA (PRP) AND PLATELET DERIVATIVES FOR TOPICAL THERAPY. WHAT IS TRUE FROM THE BIOLOGIC VIEW POINT?", ISBT SCIENCE SERIES, vol. 2, 2007, pages 272 - 281 * |
Cited By (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2016116560A1 (en) | 2015-01-21 | 2016-07-28 | Turkof Edvin | Platelet concentrate for increase of cell regeneration and cell growth |
US11564943B2 (en) | 2015-01-21 | 2023-01-31 | Edvin Turkof | Platelet concentrate for increase of cell regeneration and cell growth |
AU2015388056B2 (en) * | 2015-03-23 | 2019-04-04 | HCell Inc. | Method for promoting hair growth and preventing hair loss of patient, composition therefor, and method for preparing composition |
CN109310711A (en) * | 2016-03-23 | 2019-02-05 | 里尔大学中心医院 | The preparation of pellet platelets lysate and its purposes for the treatment of the nervous system disease |
US11083687B2 (en) * | 2016-06-03 | 2021-08-10 | Embryogenesis Pty Ltd | Skin care formulation |
WO2018015973A1 (en) | 2016-07-21 | 2018-01-25 | Khorakiwala Habil F | Topical pharmaceutical composition for promoting hair growth and/or reducing hair loss |
CN109803671A (en) * | 2016-07-21 | 2019-05-24 | 哈比尔·F·赫拉基瓦拉 | For promoting hair growth and reducing the local medicine composition of alopecia |
WO2022035849A1 (en) * | 2020-08-10 | 2022-02-17 | Venturis Therapeutics, Inc. | Methods reducing hair loss and/or increasing hair regrowth |
CN116831983A (en) * | 2023-09-02 | 2023-10-03 | 潍坊市人民医院(潍坊市公共卫生临床中心) | Concentrated growth factor composite gel preparation, preparation method and hair growth application |
CN116831983B (en) * | 2023-09-02 | 2023-12-29 | 潍坊市人民医院(潍坊市公共卫生临床中心) | Concentrated growth factor composite gel preparation, preparation method and hair growth application |
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