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WO1996009064A1 - Procede et preparation pharmaceutique pour la prevention et le traitement des lesions cerebrales - Google Patents

Procede et preparation pharmaceutique pour la prevention et le traitement des lesions cerebrales Download PDF

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Publication number
WO1996009064A1
WO1996009064A1 PCT/US1995/012057 US9512057W WO9609064A1 WO 1996009064 A1 WO1996009064 A1 WO 1996009064A1 US 9512057 W US9512057 W US 9512057W WO 9609064 A1 WO9609064 A1 WO 9609064A1
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WIPO (PCT)
Prior art keywords
pacap
compound
pacap27
composition
pacap38
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PCT/US1995/012057
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English (en)
Inventor
Akira Arimura
Original Assignee
The Administrators Of The Tulane Educational Fund
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Application filed by The Administrators Of The Tulane Educational Fund filed Critical The Administrators Of The Tulane Educational Fund
Priority to US08/809,500 priority Critical patent/US6680295B1/en
Priority to AU36810/95A priority patent/AU3681095A/en
Publication of WO1996009064A1 publication Critical patent/WO1996009064A1/fr

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/575—Hormones
    • C07K14/57563—Vasoactive intestinal peptide [VIP]; Related peptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
    • A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • the present invention relates to a method for the prevention and treatment of neuronal cell damage in the brain of mammals, including, for example, damage induced by ischemia followed by reperfusion or that induced by toxic substances.
  • Neuronal cell damage results from many causes. The most common cause of neuronal cell damage is ischemia/reperfusion of the brain following vascular occlusion by thrombosis or embolism, and cardiac failure, which occasionally occurs during heart surgery.
  • Neuronal cell damage or death can be caused both by the ischemia and by reperfusion of blood after the transient ischemia has subsided. See W. D. Dietrich, "Morphological Manifestations of Reperfusion Injury in Brain," Ann. N.Y. Acad. Sci. 723: 15 (1994). Transient ischemia of the brain sometimes also occurs in newborn babies during complicated deliveries. Neuronal cell damage or death can also be induced by various toxic substances, including, for example, the gp120 envelope glycoprotein of the HIV virus, other viral toxins, bacterial toxins, animal toxins, e.g., snake venom, toxic waste, e.g., methylated mercury, and others.
  • the methods and compositions of the present invention can be used to protect and/or attenuate neuronal cells in the brain, spinal cord or elsewhere in the body, caused by trauma, infection e.g., encephalitis, AIDS or degenerative diseases such as Parkinson's disease, Alzheimer's disease, etc.
  • PACAP Pituitary adenylate c ⁇ clase activating polypeptide
  • PACAP is a new member of the secretin/glucagon/vasoactive intestinal peptide (VIP) family, being the most homologous to VIP, but its adenylate cyclase stimulating activity in cultured pituitary cells, neurons and astrocytes is about 1 ,000 - 10,000 times greater than VIP, A. Miyata et al., supra.
  • VIP secretin/glucagon/vasoactive intestinal peptide
  • PACAP is a pleiotropic neuropeptide, exhibiting a number of neurotropic actions in different organs and tissues. For example, PACAP enhances proliferation and differentiation of sympathetic neuroblasts, stimulates neurite outgrowth of an adrenal chromaffin cell line, PC 12 cells, and stimulates growth of astrocytes, E.
  • PACAP receptors There are two types of PACAP receptors (Gottschall et al., Endocrinology 127/1 :272-277 (1990); Shivers et al., Endocrinology, 128/6:3055-3056 (1991 ); A. Arimura, Trends in Endocrinology and Metabolism, 3/8: 288-294 (1992).
  • the Type I PACAP receptor specifically binds to PACAP with high affinity, but do not bind to VIP.
  • the Type II PACAP receptors bind to both PACAP and VIP with similar high affinities, and may be very similar to or identical with the VIP receptor.
  • the present invention relates to a method and pharmaceutical preparations for treating or preventing neuronal cell damage in mammals, comprising administering a effective amount of a PACAP, or an agonist, analog or derivative thereof having PACAP neurotrophic activity, in a pharmaceutically acceptable carrier, in a concentration which is effective for protection of neuronal nerve cells in vivo.
  • the present invention involves a method of treatment of mammalian neuronal cells in which the concentration of the PACAP compound is between about 10 "15 and 10 "12 M in the tissues.
  • concentration of the PACAP compound is between about 10 "15 and 10 "12 M in the tissues.
  • the preferred concentration range for treatment with the PACAP compounds of the present invention lies between about 10 "14 and about 10 12 M and another range of concentration lies between about 10 "11 and about 10 9 M.
  • the preferred concentration range for treatment is the range between about 10 '14 and about 10 '12 M in the tissue, which permits treatment of the subject with minimal risks of side effects from the treatment. The present discovery makes possible the use of such
  • PACAP pharmaceuticals in extremely low concentrations to provide very substantial protection of neuronal cells, such as brain cells, from death due to transient ischemia, reperfusion, toxic substances, trauma, or other causes.
  • compositions in accordance with the present invention include PACAP, in either of its forms, commonly referred to as PACAP38 and PACAP27, as well as any peptide or non-peptide agonist for PACAP receptors, especially agonists for the Type I PACAP receptor.
  • PACAP PACAP
  • the PACAP compound is a polypeptide, or a salt or derivative thereof, which contains at least twelve amino acids joined in a sequence corresponding to a part of the sequence shown for PACAP38 in Figure 1 , and which binds to at least one receptor which binds to PACAP.
  • PACAP 12 agonist refers to a polypeptide, or salt or derivative thereof, which has at least 12 amino acids corresponding in sequence to some part of the amino acid sequence of PACAP38, as shown in Figure 1 , and which binds to at least one PACAP receptor.
  • PACAP 23 agonist and PACAP 27 agonist refer to pol ⁇ peptides, or salts or derivatives thereof, which has at least 23 and 27 amino acids, respectively, corresponding in sequence to some part of the amino acid sequence of PACAP38, as shown in Figure 1 , and which binds to at least one PACAP receptor.
  • Determination of the amino acid sequence of the polypeptide, and determination as to whether it binds to a PACAP receptor, are both well within the skill in the art.
  • the ability to treat neuronal cells at such low concentrations also makes possible the administration of the PACAP compounds intravenously or otherwise into the blood, in concentrations sufficient to provide PACAP compound transfers across the blood/brain barrier sufficient to provide concentrations of the PACAP compound in contact with the neuronal cells which are effective to protect and/or resuscitate the traumatized neuronal cells.
  • PACAP compound is effective in protecting and/or resuscitating traumatized neuron cells up to at least 24 hours after injury.
  • PACAP compound has the general formula:
  • X-PACAP (a . bl -Y wherein X is H or a solubility effecting group such as a C,. 20 carboxylic acid moiety, such as formyl, acetyl, etc.; a and b are N and C terminus amino acids taken in the sequence of PACAP38 as shown in Fig. 1 , and Y is H, NH 2 , OH, or C 1-4 carboxy.
  • X is a fatty acid moiety, preferably derived from Laurie, Myristic, Palmitic, Stearic, or Oleic acid, most preferably from Palmitic or Stearic acid.
  • PACAP38 is PACAP ⁇ . 38
  • the polypeptide can be substituted at either end with moieties which favorably effect the solubility in the carrier, or favorably effect the ability of the PACAP compound to transfer across the blood brain barrier without substantially adversely effecting the effectiveness of the compound.
  • X can be an organic acid or salt thereof, preferably containing only alkyl groups of C, ⁇ , preferably C,. 20 , or a residue from such an acid, e.g., an ether derived from such an acid.
  • Low molecular weight (C 4 ) acids or acid residues can be used to increase the solubility of the polypeptide in the pharmaceutical composition, or in bodily fluids.
  • Larger molecular weight moieties such as the C 12 . 2 o - on 9 chain fatty acid residues, can be used to enhance the transferability of the PACAP compound across the blood brain/barrier.
  • Substituents at the C- terminus of the polypeptide can also be used to enhance the solubility of the PACAP compound without deleteriously effecting its usefulness.
  • the amino (NH 2 ) group on the C-terminal amino acid can be substituted by a hydroxyl group or a lower (C,. 4 ) alcohol or carboxyl group.
  • substitutions of certain of the amino acids in the PACAP sequence it is possible to make various substitutions of certain of the amino acids in the PACAP sequence, to make minor adjustments in the physical properties of the molecule without substantially effecting its usefulness in treatment of neuronal cells.
  • substitution of less reactive amino acids can provide increased stability and shelf life of the pharmaceutical composition.
  • substitutions it is possible to make one or more of the following substitutions:
  • [Glu 3,8 ]PACAP. 1 . 27) -NH 2 refer to PACAP27 wherein the asparagine at position 3 and the asparagine at position 8 have each been replaced by glutamic acid.
  • PACAP 38 The most preferred active ingredient of the pharmaceutical composition is PACAP 38, its salts and derivatives.
  • PACAP27 the last most preferred is PACAP27, its salts and derivatives.
  • PACAP27 the polypeptides which have the same amino acid sequence as amino acids 1 -27 and 1 -38, respectively, of PACAP38, as shown in Figure 1 .
  • PACAP38 and PACAP27 come with the scope of the terms “PACAP 12 " "PACAP 23 " and "PACAP 27 "
  • PACAP type compounds include:
  • N°-Acetyl-PACAP 1 27 -NH 2 1 1 .
  • N ⁇ X-PACAP ⁇ a ⁇ -NH j , X C 10 . 18 fatty acid 41 .
  • N ⁇ -X-PACAP 2 . 38 -NH 2 , X C 1f 18 fatty acid
  • PACAP receptor 1 Any peptide or non-peptide agonist (except those listed above) for PACAP receptor 1 and organic and inorganic salts thereof.
  • PACAP27 SEQ ID NO:2.
  • Fig. 2. is a representative displacement curve of [ 126 I]PACAP27 binding to rat brain membrane preparations by PACAP and structurally related peptides. Inset shows Scatchard plot analysis of the data from [ 125 I]PACAP27 binding displaced by unlabeled PACAP27.
  • Fig. 5. shows the number of dead cells in the dorsomedial population of dorsal root ganglia (DRG18) in the chick embryo on day E9. The control group received vehicle and the experimental group received PACAP38 (25nM in the egg) daily from E3 1/2 to E8 1/2. Number is the mean of the numbers of degenerating cells in 6 ganglia in each group. Bar indicates SEM.
  • Fig. 6. shows the number of viable motoneurons in the lateral motor column (LMC) of chick embryos on E10. Embryos received vehicle, or 5 nM or 25nM PACAP (concentrations in the egg after injection) daily from E6 to E9. Average LMC is represented by 320-340 sections and cell counts were made on every tenth section. Sum of live cells in all counted sections multiplied by 10 is shown in this figure. Numbers indicate mean numbers of live cells in 6 LMC in each group. Bars indicate SME.
  • LMC lateral motor column
  • Fig. 7. shows distribution of motoneurons across LMC in chick embryos at E10. Mean of motoneuron numbers per section of 1 -9 segments of lumbar column of vehicle (open circle) and 25 nM PACAP treated (close circle) embryos. Note the difference is obvious only in the middle and mid-caudal segments.
  • Fig. 8. shows the comparison of PACAP38 and PACAP27: prevention of gp120-induced neuronal cell death in dissociated hippocampal cultures.
  • the cultures were prepared from 16-17 day-old mice embryos (C57B1 /6) as previously described. Brenneman, et al., Nature 335: 639-642 (1988). Before treatment, the cultures were given a complete change of medium. One week old cultures were treated for five days with 1 pM gp1 20 (RFII isolate) and either PACAP38 (closed circles) or PACAP27 (open circles). Cell counts were conducted on 60 fields, 0.12 mm 2 each. Neurons were identified immunocytochemicall ⁇ using antiserum against neuron specific enolase.
  • Fig. 9. shows antiserum to PACAP27 produced neuronal cell death in dissociated hippocampal cultures: prevention by PACAP27.
  • Antiserum was obtained from rabbits as described previously. Arimura, et al., Endocrinology 129: 2787-2789 ( 1 991 ). A comparison was made between cultures treated with antiserum alone (closed circles) and dishes given antiserum plus 0.1 nM PACAP27 (open circles). The experimental conditions were the same as that described in Figure 7. Each value is the mean of 3-5 dishes. The error bar is the standard error. A significant decrease from control was observed at 1 : 100 antiserum dilution (P ⁇ 0.001 ).
  • FIGs. 10A and 10B are graphs showing cAMP response to PACAP in cultured rat neurons. Neuron cultures were prepared from brains of 1 da ⁇ -old rats. 1 x 10 7 cells were plated in each dish (35 mm diameter). Neurons were cultured for two weeks before experiment. cAMP in cells were determined by RIA various time intervals after addition of PACAP.
  • Figure 10A shows temporal pattern of intracellular cAMP concentrations after addition of 10 '9 M PACAP38.
  • Figure 10B shows the mean of intracellular cAMP concentrations determined 10 min after addition of varying doses of PACAP38, PACAP27 or VIP. ' indicates level of significance, P ⁇ 0.05.
  • Figs. 1 1A and 1 1 B are graphs showing cAMP response of cultured rat astrocytes to PACAP.
  • Astrocytes cultures were prepared from brain of neonatal rats. Astrocytes were allowed to proliferate for 7-10 days before experiments.
  • Figure 1 1 A shows temporal pattern of intracellular cAMP concentrations after addition of 10 "9 M PACAP38.
  • Figure 1 1 B shows the mean of intracellular cAMP concentrations determined 10 min after addition of varying doses of PACAP38, PACAP27 or VIP. ' indicates level of significance, P ⁇ 0.05.
  • Other experimental protocols are same as shown in Fig. 10.
  • Fig. 12 is a graph showing the prevention of ischemia induced neuronal cell death in rat hippocampus using PACAP38.
  • Each column indicates viable cell number 7 days after 15 min ischemia of the forebrain of rats which were intracerebroventricularly administered the vehicle alone or the vehicle containing varying amounts of PACAP38.
  • the numbers of cells are expressed by percentages of live cells as compared with the cells in the same area of the intact rat hippocampus.
  • a bolus administration of 450 ng PACAP 38/10 ⁇ H 2 O over 10 min followed by continuous infusion of the peptide at 45 ng/ ⁇ l/hr over seven days was the most effective in cytoprotection. The higher dose was less effective.
  • Figs. 13A-F are microphotographs of coronal sections of the right hippocampal CA1 from intact rats and rats subjected to forebrain ischemia for 15 min. The sections were stained with cresyl violet (A, B, C) and immunostained for MAPII (D, E, F).
  • A, D Intact rats.
  • B, E 7 days after ischemia in vehicle-infused (icv) rats. Note severe damage to pyramidal cells in CA1 (B), and marked decrease in MAPII immunoreactivity in the denditric fields of the CA1 area (E).
  • C, F 7 days after ischemia in rats infused with PACAP38 icv by an implanted osmotic pump over one week at 1 pmol/h.
  • PACAP38 administration started immediately after ischemia.
  • Fig. 14. is a graph similar to Fig. 12, showing the prevention of ischemia- induced neuronal cell death in rat hippocampus using PACAP38 as described in Example 6.
  • Fig. 15 is a graph showing 25 l- PACAP 38 levels in brain tissue 10 minutes after icv injection, at 1 X 10 6 cpm 1 5 l- PACAP38.
  • Fig. 16 is a graph showing the effects of icv infusion of PACAP38 into the right lateral cerebroventricle on ischemia-induced neuronal cell death in the hippocampal CA1 in rats.
  • the ordinate shows the mean number _+_ SE of viable neurons per reticle (220 ⁇ m) 7 days after ischemia.
  • Fig. 17 is a graph showing the cytoprotective effect of iv infusion of PACAP38 which started immediately after ischemia in rats. Both 160 pmol/hr and 16 pmol/hr PACAP38 significantly attenuated ischemia-induced neuronal cell death in CA1 7 days after ischemia. The small dose of PACAP38 appeared to exhibit greater effect.
  • Fig. 18 is a graph showing the cytoprotective effect of iv infusion of PACAP38 which started 24 hrs. after ischemia in rats. Both 16 pmol/hr and 160 pmol/hr PACAP38 significantly attenuated ischemia- induced neuronal cell death in CA1 7 days after ischemia.
  • PACAP sustained intracerebroventricular administration of PACAP can be used to protect neuronal cells and significantly prevent neuronal cell damage and death induced by ischemia, hemorrhage, trauma, toxic substances or other causes. It is particularly effective in treating neuronal cell damage caused by transient ischemia followed by reperfusion. Unexpectedly, the present inventors found that PACAP compounds are most effective in vivo in a particular concentration range, above which the effectiveness drops off.
  • the present invention provides a method for the treatment and prevention of neuronal cell damage and death, preferably brain cell damage and death, in vitro and/or in vivo, induced by various causes comprising administrating to neuronal cells or to a mammal in need thereof an effective amount of a PACAP or an agonist thereof, which interacts with specific Type I PACAP receptors.
  • the in vivo mode of administration is intraventricular or intravenous.
  • PACAP neurotrophic action
  • PACAP- stimulates proliferation of sympathetic neuroblast and neurite outgrowth in PC12 cells directly, but it requires nanomolar concentration.
  • Cytoprotective action of PACAP on chick embryos also requires nanomolar concentration of the peptide, much higher than the effective concentrations discovered in connection with the present invention. That type of cytoprotective action is believed to be mediated via or through adenylate cyclase activation by PACAP.
  • Applicant has also shown that when neuronal cells are plated on a feeder layer of astrocytes, addition of HIV envelope glycoprotein gp120 results in significant cell death. Yet, when small concentration of PACAP, e.g., 10 "13 M, was added to the culture, the gp120-induced neuronal cell death was completely prevented. In such cases, higher concentrations of PACAP have been shown to be less effective. Since this cytoprotective action is difficult to demonstrate in neuron cultures in the absence of astrocytes, this cytoprotective action is considered to be mediated through astrocytes, which express an extremely high affinity Type I PACAP receptors.
  • PACAP at such a low concentration does not stimulate adenylate cyclase or phospholipase C, and as a result, the cytoprotective action may be mediated by one or more other second messengers, most likely by alteration of intracellular Ca 2+ .
  • PACAP which has been found to increase intracellular Ca 2 + of cultured pancreatic beta cells at 10 '13 M (Yada et al., J. Biol. Chem. , in press 1994). Based on this evidence it is believed that a subtype of Type I PACAP receptors with an extremely high affinity may be expressed on astrocytes in vivo under certain conditions, such as ischemia-induced brain injury.
  • astrocytes with Type I PACAP receptors begin appearing 2 days after ischemia and the number increases over 7 days after ischemia.
  • Applicants' in vitro examination of gp120-induced brain cell death suggests that the interaction of PACAP with these newly expressed receptors in astrocytes may stimulate synthesis and release of a neurosurvival factor that prevents neuronal cell death.
  • PACAP may be used in the treatment and prevention of neuronal cell damage resulting from ischemia/reperfusion, trauma, hemorrhage, infection and exposure to toxic substances.
  • PACAP can also be used for 1 ) treatment of congestive heart failure of neonate, through its inotropic action and stimulation of adrenalin secretion in vivo; 2) treatment of neuropathy, such as diabetic neuropathy; 3) treatment of spinal cord injury; 4) treatment of ischemia/reperfusion induced lung injury (cAMP has been shown to prevent such injury, and PACAP is a potent stimulater for cAMP production); 5) treatment of ischemia/reperfusion induced cardiac injury; 6) treatment of gastric and intestinal ulcer (PACAP regulates production of various growth factors which are known to prevent ulcer); 7) stimulation of neonatal and prenatal brain development; 8) protection of transplanted neural cells in the brain; 9) treatment of certain male infertility; 10) improvement of brain circulation; 1 1 ) treatment of shock, such as the condition
  • PACAP stimulates growth and differentiation of neuronal and glial cells and cell lines in vitro. Any cell line which possesses PACAP receptors can be stimulated by PACAP in growth and differentiation.
  • PACAP38, PACAP27, PACAP their agonist analogs, precursors and salts are prepared in a manner which will be apparent to the skilled in the art.
  • the peptides were synthesized by solid phase techniques using an automated peptide synthesizer (Beckman 990B). 4-methyl benzhydrylamine resin and PAM-resin were employed for the synthese of C-terminal amide form peptides and C-terminal free form peptides, respectively.
  • N"-Boc-amino acid derivatives such as: Boc-Lys(CI-Z)-OH, Boc-Asn-OH, Boc-Val-OH, Boc-Arg(Tos)- OH, Boc-Gin-OH, Boc-Tyr(Br-Z)-OH, Boc-Leu-OH, Boc-Ala- OH, Boc-Met-OH, Boc-Ser(Bzl)-OH, Boc-Asp(OBzl)-OH, Boc- Thr(Bzl)-OH, Boc-Phe-OH, Boc-lle-OH, Boc-His(Tos)-OH, Boc-Glu(OBzl)-OH and Boc-Nle-OH
  • N°Boc amino acid derivatives were successively introduced to the peptide chain in the presence of diisopropylcarbodiimide in dichloromethane with the exception of Boc-Asn-OH and Boc-Glyn-OH which were coupled in the presence of 1 -hydroxybenzotriazole as a catalyst in DMF.
  • the completed, protected peptide resins (90.025 mmol each) were treated with 20 mL of anhydrous hydrogen fluoride containing 10% anisole and 100 mg of dithiothreitol for 45 min at 0°C. After removal of the hydrogen fluoride under a stream of nitrogen, the free peptides were precipitated with either ether or ethyl acetate, filtered, and extracted with 2M AcOH.
  • the crude peptides were obtained.
  • the crude peptides were purified by gel filtration on a column of SEPHADEX G-50 fine (2.5 x 100cm) using 2M AcOH containing 0.02% ?-mercaptoethanol as an eluent, followed by preparative reverse phase HPLC column (1.5 x 50 cm) of Vydac C-18 silica (15-20 mm particle size), which was eluted with a linear gradient of 10-35% acetonitrile in 0.1 % TFA at a flow rate of 3mL/min.
  • PACAP or its agonists may be intracerebroventricularly administrated to a host in need thereof utilizing a variety of means known to the skilled artisan including, for example, an infusion system such as The SynchromMed Infusion System with Catheter Access Port (Medtronic Neurological, Minneapolis, MN).
  • Suitable compositions for direct administration to the brain include the polypeptide in a carrier such as spinal fluid, artificial spinal fluid, or a combination of physiological saline, Ringer's solution, glucose (e.g., 3-7%, preferably about 5% by weight), and an isotonic phosphate buffer (pH of about 7).
  • Artificial cerebrospinal fluid comprises about 1 28 mM NaCI, 2.6 mM KCI, 1 .3 mM CaCI 2 , 20 mM NaHCO 3 , 1 .3 mM Na 2 HPO4, pH 7.35 and contains 0.1 % bovine serum albumin.
  • 0.9% saline containing 0.1 % bovine serum albumin may be used.
  • Bovine serum albumin is used for protection of loss of the peptide due to adsorption.
  • Bovine serum albumin can be replaced by any other inert protein such as human serum albumin and gelatin.
  • the amount of PACAP compound to be administered is sufficient to achieve a concentration in the tissue to be treated of from about 10 '15 to about 10 12 M, more preferably from about 10 '14 to about 10 12 M.
  • a concentration in the tissue to be treated of from about 10 '15 to about 10 12 M, more preferably from about 10 '14 to about 10 12 M.
  • icv intracerebroventricular
  • the dosage ranges corresponding to the above concentration ranges are from about 0.1 6 ng/kg body weight/hour to 160 ng/kg body weight/hour, preferably either from 1.6 ng/kg body weight/hour to 160 ng/kg body weight/hour.
  • a 300 g for a 300 g.
  • the optimal dosage for icv infusion was about 1 pmol/ ⁇ l/hr for a seven day course of treatment, which is equal to a rate of 3.33 pmol (15.7 ng)/3.33 ⁇ l/kg/h.
  • the infusion rate is about 233 pmol (1.1 ⁇ g)/233 ⁇ l/h.
  • the concentration of PACAP is 1 nmol/ml (4.5 g/ml) for icv infusion.
  • An infusion system such as a SynchroMed Infusion System can deliver solution at a range between 4 ⁇ and 900 ⁇ l/h.
  • An infusion rate of 50 - 500 ⁇ l is preferred.
  • Suitable concentrations of the PACAP compound in the pharmaceutically acceptable carrier for icv infusion are prepared according to the infusion rate. For example, when one wishes to infuse 1.1 ⁇ g/hour and an infusion rate of 50 l/h is chosen, the concentration of PACAP is 1.1 ⁇ g/50 ⁇ or 22 g/ml. When 500 ⁇ l/h is chosen, a 2.2 //g/ml solution is prepared. Accordingly, suitable concentrations of the PACAP compound in pharmaceutically acceptable carrier for icv infusion at a rate of 1.1 ⁇ g/hour are from about 1.2 /g/ml to about 300 /yg/ml, preferably from about 2.2 ⁇ g/ml to about 22 ⁇ g/ml. Suitable concentration ranges for other infusion rates can be likewise calculated.
  • PACAP administered icv is not evenly distributed throughout the brain tissues.
  • administration of 125 l labelled PACAP into the right lateral ventricle resulted in four times greater CPM in the right hippocampus than in the left.
  • the amount of PACAP administered must be adjusted accordingly.
  • the administration of PACAP is not limited to the intracerebroventricular route. Despite the existence of blood-brain- barrier, a small portion of intravenously administered PACAP is transported into the brain. Banks, et al., Journal of Pharmacology and Experimental Therapeutics, 267: No. 2 690-696 (1993).
  • PACAP38 is transported into the brain in a saturated manner. Therefore, it is also possible to administer PACAP and its analogs by a prolonged intravenous or even subcutaneous infusion to attain the optimal concentration of PACAP in the brain tissues for attaining the similar cytoprotective effect.
  • the percentage of the amount of circulating PACAP is transported into the brain in a saturated manner. Therefore, it is also possible to administer PACAP and its analogs by a prolonged intravenous or even subcutaneous infusion to attain the optimal concentration of PACAP in the brain tissues for attaining the similar cytoprotective effect. The percentage of the amount of circulating
  • PACAP which enters into the brain can be precisely calculated.
  • Analogs of PACAP with a greater lipophilic nature such as PACAP analogs with fatty acid residue at the N-terminus, for example: N ⁇ stearyl-INIe ⁇ lPACAP L - j ⁇ -NH j or N ⁇ -stearyl-[Nle 17 ]PACAP 2 38 - NH 2 may be more preferable for such modes of administration.
  • compositions for intravenous administration are preferably prepared by forming a solution of the PACAP compound in a water- soluble solvent (e.g. physiological saline, Ringer's solution).
  • a water- soluble solvent e.g. physiological saline, Ringer's solution.
  • the dosage ranges corresponding to the above concentration ranges from about .25 ng/kg body weight/hour to 25 ⁇ g/kg body weight/hour, preferably from .25 ng/kg body weight/hour to about 2.5 ⁇ g/kg body weight/hour, most preferably from about 2.5 ng/kg body weight/hour to about 250 ng/kg body weight/hour. Based on the above a human patient weighing 70 kg, the optimal IV dose would be about 1.8 ⁇ g/h.
  • solution can be infused at, for example, 100 ⁇ l/h, and thus, 1.8 ⁇ g PACAP is dissolved in 100 ⁇ l to make 18 ⁇ g/ml in 0.9% saline, 5% glucose solution or a solution for IV infusion which contains various salts (these solutions are commercially available and used to meet patients' conditions.)
  • Infusion rates can be modified to a rate faster or slower than 100 ⁇ l/h, and the concentrations of PACAP should be adjusted accordingly. For example, when the infusion rate is 1000 ⁇ l/h, the concentration of PACAP is 1.8 ⁇ g/ml.
  • the amount of PACAP in a pharmaceutical composition for intravenous administration is 10 to 100,000 times the amount that is effective at the active region, preferably 100 to 10,000 times and most preferably 500 to 5,000 times.
  • additives such as a dissolution aid (e.g. sodium salicylate, sodium acetate), buffer (e.g. sodium citrate, glycerine), isotonizing agent (e.g. glucose, invert sugar), stabilizer (e.g. human serum albumin, polyethylene glycol), preservatives (e.g. benzyl alcohol, phenol), or analgesics (e.g. benzalkonium chloride, procaine hydrochloride).
  • a dissolution aid e.g. sodium salicylate, sodium acetate
  • buffer e.g. sodium citrate, glycerine
  • isotonizing agent e.g. glucose, invert sugar
  • stabilizer e.g. human serum albumin, polyethylene glycol
  • preservatives e.g. benzyl alcohol, phenol
  • analgesics e.g. benzalkonium chloride, procaine hydrochloride
  • compositions for administration of PACAP include those suitable for oral, rectal, nasal, topical (including buccal and sublingual), vaginal or parenteral (including subcutaneous, intramuscular, intravenous and intradermal) administration.
  • the formulations may conveniently be presented in unit dosage form, e.g., tablets and sustained release capsules, and in liposomes and may be prepared by any methods well known in the art of pharmacy.
  • Such methods include the step of bringing into association the ingredients to be administered with the carrier which constitutes one or more accessory ingredients.
  • the compositions are prepared by uniformly and intimately bringing into association the active ingredients with liquid carriers, liposomes or finely divided solid carriers or both, and then if necessary shaping the product.
  • the amount of PACAP in a composition for parenteral administration is 100 to 1 ,0000,000 times, the amount that is effective at the active region, preferably 1 ,000 to 100,000 times and most preferably 5,000 to 50,000 times.
  • compositions of the present invention suitable for oral administration may be presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or a suspension in an aqueous liquid or a non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion or packed in liposomes and as a bolus, etc.
  • a tablet may be made by compression or molding, optionally with one or more accessory ingredients.
  • Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with a binder, lubricant, inert diluent, preservative, surface-active or dispersing water.
  • Molded tablets may be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent.
  • the tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein.
  • compositions suitable for topical administration to the skin may be presented as ointments, creams, gels and pastes comprising the compound and a pharmaceutically acceptable carrier.
  • a suitable topical delivery system is a transdermal patch containing the ingredient to be administered.
  • Sublingual tablets can be prepared by using binders (e.g. hydroxypropylcellulose, hydroxypropylmethylcellulose, polyethylene gl ⁇ col), disintegrating agent (e.g. starch, carboxymeth ⁇ lcellulose calcium,), lubricant (e.g. magnesium stearate, talc)
  • binders e.g. hydroxypropylcellulose, hydroxypropylmethylcellulose, polyethylene gl ⁇ col
  • disintegrating agent e.g. starch, carboxymeth ⁇ lcellulose calcium,
  • lubricant e.g. magnesium stearate, talc
  • compositions suitable for rectal administration may be presented as a suppository with a suitable base comprising, for example, polyethylene glycol 600, cocoa butter or a salicylate.
  • compositions suitable for nasal administration wherein the carrier is a solid include a coarse powder having a particle size, for example, in the range 20 to 500 microns.
  • Suitable formulations wherein the carrier is a liquid, for administration, as for example, a nasal spray or as nasal drops, include aqueous or oily solutions of the active ingredient.
  • Compositions suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams or spray formulations containing in addition to the active ingredient such carriers as are known in the art to be appropriate.
  • compositions suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents.
  • the formulations may be presented in unit-dose or multi-dose containers, for example, sealed ampules and vials, and may be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example, water for injections, immediately prior to use.
  • Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tables of the kind previously described.
  • formulations of this invention may include other agents conventional in the art having regard to the type of formulation in question, for example, those suitable for oral administration may include flavoring agents.
  • compositions comprising these compounds are extremely low in toxicity.
  • the present invention is further illustrated by the following examples. These examples are provided to aid in the understanding of the invention and are not construed as a limitation thereof.
  • mice Male CD rats (Charles River Breeding Labs., Wilmington, MA) were used throughout this example. The following regions of the brain were dissected at the indicated ages: embryonic day E14 (whole brain); E18 (whole brain, neocortex, hippocampus, and diencephalon); postnatal days P0, P7 and P14 (whole brain, neocortex, hippocampus, diencephalon, and cerebellum); and 1 -, 2- and 3-months old (MO) (whole brain, neocortex, hippocampus, diencephalon, and cerebellum). After dissection, each region was immediately frozen on dry-ice and stored at -70C for up to 2 months until the receptor and peptide levels were determined.
  • E14 embryonic day E14
  • E18 whole brain, neocortex, hippocampus, and diencephalon
  • postnatal days P0, P7 and P14 whole brain, neocortex, hippocampus, dience
  • the Kd and Bmax for PACAP27 binding were 0.809 _+ 0.086 nM and 7.83 _ ⁇ _ 2.08 pmol/mg protein respectively.
  • VIP, GHRH, glucagon and secretin did not displace 125 I-PACAP27 binding.
  • PACAP38 displaced 25 I-PACAP27 binding more effectively than did PACAP27, but the displacement slope with unlabeled PACAP38 was parallel to that with unlabeled PACAP27.
  • Scattered plot analysis of the displacement curve for PACAP 38 also indicated the existence of a single class of binding sites.
  • PACAP38 had a significantly higher (about 10 fold) affinity for the receptor than PACAP27, there was no significant difference in Bmax between PACAP27 and PACAP38, suggesting that these peptides share the same binding site but with different affinities. Therefore, additional Scatchard plot analysis was performed analyzing both displacement curves for unlabeled PACAP27 and PACAP38 simultaneously assuming that PACAP27 and PACAP38 share the same binding site. There were no significant differences in Kd and Bmax estimated by these 3 analyses. The Kds for PACAP27 and PACAP38 in the E1 8 brain membrane preparations were comparable to those in membrane preparations from adult rat hypothalamus and pituitary.
  • PACAP38 is immunoreactivity determined by RIA was detected
  • PACAP38 (2.1 _ ⁇ _ 1.1 pg/mg wet tissue) in the whole brain extracts at E14.
  • the embryos were weighed and staged by measuring the toe length. Hamburger, et al., Journal of Morphology 88: 49-92. It should be noted that the experiment was designed so that the treatment began after the period of extensive neural proliferation in the embryo. This ensured that any increase in cell number was due to a prevention of natural cell death, not a stimulation of mitoses.
  • tissue between the cervical and lumbar regions of the spinal cord was removed and placed in Carnoy's fixative for two hours. Once fixed, the tissues were transferred directly to a thionine solution and stained en bloc for 15 hrs. The blocks were subjected to serial dehydration in graded alcohol and embedded in paraffin. Tissues were sectioned el at 10 m, mounted on gelatin coated slides, deparaffinized, and cover slipped with Cytoseal.
  • the preparation of the lumbar column of the spinal cord was identical to that for the DRG 18 study, with the following exceptions;
  • the entire lumbar spinal column (segments 21 -32) was removed, dissected free of skin, fixed in Carnoy's solution, stained, embedded in paraffin, serial sectioned, and mounted as before.
  • Both the control and experimental groups were administered vehicle or PACAP38 respectively, daily from E6 through E9 using the same technique described above, and sacrificed on E10 (Stage 36).
  • the lower brachial and upper thoracic ganglia were reconstructed for each embryo. Reconstruction was achieved by marking the block, slide, row, and section numbers which corresponded to the first and last sections of the ganglia in each bloc.
  • the brachial plexus was used as the primary landmark for DRG 16 and the third ganglia from DRG 16 was identified as DRG 18.
  • the LMC was reconstructed in a similar manner, by citing the bloc, slide, row and section numbers.
  • the motor column stained very heavily compared to the rest of the spinal cord.
  • the magnification used (x400), one can observe DRG18 divided into two populations of cells: large differentiating ventrolateral (VL) and small, late differentiating dorsomedial (DM) cells. Although the boundary between the two populations of neurons was sometimes indistinct, it was possible to identify individual cells based upon their morphologies.
  • VL ventrolateral
  • DM dorsomedial
  • the number of degenerating cells increased sharply beginning at E4 1 /2 (Stage 25) peaked at Stage 27 and then declined immediately. Hamburger, et al., Journal of Neuroendocrinology 1 :60-71 (1981 ).
  • DM population a rapid increase in the number of degenerating cells began at stage 33; thus the degeneration periods for the VL and DM cells barely overlap.
  • DM degeneration The peak of DM degeneration was reached within half a day, at stage 34, and a sharp decline began at stage 35.
  • DM population of neurons was studied. Degenerating cells displayed varying morphologies from large vacuolated cells to mere fragments. However, most degenerating neurons appeared as deeply stained, homogenous spheres. Macrophages containing debris of several cells were encountered rarely and counted as two degenerating cells. Every second section of the ganglia was counted, and the sum multiplied by two to obtain a data point. A total of 6 ganglia were counted in each of the control and experimental groups, respectively.
  • LMC lumbar motor column
  • PACAP38 25 nM in the egg
  • the number of the degenerating cells in the PACAP38-treated embryos was 37.2% less than that in the untreated embryos. The difference was highly significant (p ⁇ 0.01 ).
  • PACAP38 increased the survival of both sensory and motoneurons during the embryonic development of chick embryo.
  • chick embryos at stage 23-34 express the receptors for PACAP38.
  • Tatsuno, et al., Peptides 15: 55-60 PACAP and its receptors appear to be expressed in chick embryos.
  • the primary structure of chick PACAP is different from PACAP of mammals including rat, sheep and humans by only one amino acid (Yasuhara, et al., Reg. Peptides 37: 326 (1992), suggesting that the structure of PACAP is well conserved throughout phylogeny, and thus may play an important role for survival of neurons of animals and humans.
  • AIDS acquired immunodeficiency syndrome
  • H ⁇ V human immunodeficiency virus
  • a leading candidate for a cytotoxic product of HIV is the major external envelop protein, gp1 20.
  • Neuronal cell death associated with gp120 treatment of murine hippocampal cultures has been shown to be potently and completely prevented by VIP (Brenneman, et al., Nature 335: 639-642 (1988)) and by peptide T. Brenneman, et al., Drug Develop. Res. 15: 361-369 (1988).
  • both PACAP forms can elicit a c ⁇ toprotective action similar to that of VIP, although the bimodal response suggests a more complicated mechanism, probabl ⁇ involving more than one t ⁇ pe of receptor.
  • VIP will bind to a T ⁇ pe I PACAP receptor at an affinit ⁇ that is 1000 times less than that of PACAP; in contrast, PACAP binds to T ⁇ pe II PACAP receptors with similar affinit ⁇ as VIP. Arimura, et al., Trends Endocr. Metab. 3: 288- 294 (1 993). Arimura et al., Ann NY Acad. Sci. 739:228-243 (1994). It is possible that the PACAP-induced c ⁇ toprotective action observed at 0.1 nM ma ⁇ occur through T ⁇ pe II PACAP or VIP receptors and the activit ⁇ seen at 0.1 pM ma ⁇ result from an interaction through T ⁇ pe I
  • PACAP receptor Regardless of the phenot ⁇ pe of the receptors, PACAP like VIP increases the survival of developing hippocampal neurons exposed to lethal concentrations of the HIV envelope protein.
  • PACAP peptides have a c ⁇ toprotective effect from gp1 20
  • possibilit ⁇ neutralizing antiserum against PACAP27 (Koves, et al., Neuroendocrinology 54: 1 59-169 ( 1 991 ) and Arimura, et al.,
  • Antiserum at various dilutions was added to the cultures with or without the presence of 0.1 nM PACAP27. Addition of the antiserum alone reduced the number of neurons at 1 : 100 dilution (Fig. 9). The cell loss associated with the antiserum was prevented b ⁇ co-treatment with PACAP27. Treatment with PACAP27 alone produced a small increase in cell counts in comparison to control cultures (Fig. 9) .
  • both VIP and PACAP are recognized to stimulate the formation of cAMP (Arimura, et al., Regul. Peptides 37: 287-303 (1992) and Gozes, et al., Mol. Neurobiol 3:21 ( 1989)), the second messenger mediating the c ⁇ toprotective and survival-promoting actions of VIP and PACAP does not necessaril ⁇ utilize this signal transduction pathwa ⁇ .
  • Both PACAP38 and PACAP27 elevate intra- and extracellular cAMP in a dose-related manner in a range of 10 pM to 10nM in rat neurons and astroc ⁇ te cultures (Figs. 10 and 1 1 ).
  • the neuronal cultures showed a more rapid cAMP response to PACAP than the astroc ⁇ te cultures, but the magnitude of the intracellular accumulation of cAMP in astroc ⁇ te cultures (expressed in pmol/mg cellular protein) was about 14 times greater than the response in neurons.
  • VIP also increased cellular cAMP in both neuron and astroc ⁇ te cultures, but the activit ⁇ was approximatel ⁇ 1000 times less potent than PACAP.
  • Importantl ⁇ , no increases in cAMP for an ⁇ CNS culture s ⁇ stem has been observed at the concentrations of peptide that increase neuronal survival: 0.1 pM PACAP and 0.1 nM VIP.
  • PACAP has also been shown to increase intracellular calcium in pancreatic .-cells at 0.01 -0.1 pM. Yada, et al., J. Bio/. Chem. 269: 1 290-1 293 (1 994). We would speculate that the increase in intracellular calcium produced b ⁇ these peptides pla ⁇ s a primar ⁇ role in the c ⁇ toprotective and survival-promoting action of both PACAP and VIP.
  • the bilateral vertebrate arteries were permanentl ⁇ occluded at the first cervical vertebra b ⁇ electrocoagulation.
  • Bilateral carotid arteries were exposed and a loop of silicon rubber tubing was placed around each carotid arter ⁇ , leaving the end of the tubing outside the suture.
  • the animal was left in the laborator ⁇ with water, but without food.
  • both loops around the carotid arteries were pulled and arter ⁇ clips were applied on the loops at the portion close to the skin so that both carotid arteries were occluded and ischemia of the brain occurred as long as the clips remained.
  • the clips were removed 15 minutes later.
  • bod ⁇ temperature was monitored b ⁇ thermometer placed in the rectum and the bod ⁇ temperature was maintained at 37 °C b ⁇ radiation heat from light bulb.
  • Onl ⁇ in the animals which became unconscious was effective ischemia considered to have occurred.
  • Reperfusion of bilateral carotid arteries was assured b ⁇ direct visual examination. The incision wound was closed.
  • This example includes data of the effect of intracerebroventricular (icv) administration of PACAP38 on the neuronal cell death in the hippocampus induced b ⁇ ischemia-reperfusion in rats.
  • icv intracerebroventricular
  • e.g. 1 , 0.1 and 0.01 pmol/h over 7 da ⁇ s were tested.
  • 1 pmol/h was the most effective, though the difference from 10 pmol/h is statisticall ⁇ insignificant.
  • the range of concentrations of PACAP in the brain tissue after Intracerbroventricular (icv) infusion of the peptide was calculated. This calculation was based on the transport of 125 l-labelled peptide given icv as a bolus but slow injection over 5 min. 1 x 10 6 cpm 125 I-PACAP38 was injected icv in anesthetized rats, the cpm in the brain tissues 10 min after the injection is shown in Figure 1 5.
  • the labelled peptide was injected into the right lateral ventricle.
  • PACAP38 was icv infused into the right lateral cerebroventricle of rats having neuronal cell damage in the CA1 of the hippocampus as discussed in example 5.
  • the effect of the icv infusion of PACAP38 on ischemia-induced neuronal cell death in the hippocampal CA1 in rats was measured. The results are shown in fig. 1 6.
  • the ordinate shows the mean number _+_ SE of viable neurons per reticle (220 ⁇ m) 7 da ⁇ s after ischemia. Vehicle-treated rats showed marked neuronal cell death in both right and left hippocampus. ICV infusion of PACAP38 over one week significantl ⁇ prevented the neuronal cell death.
  • PACAP38 was administered i.v. to rats (see example 5) over one week, starting immediatel ⁇ after ischemia/reperfusion. As shown in Fig. 17, i.v. administration of the peptide significantl ⁇ attenuated neuronal cell death induced b ⁇ forebrain ischemia. The magnitude of the c ⁇ toprotective effect following iv administration of PACAP38 was ver ⁇ similar to that for icv administration (compare Fig. 16 and Fig. 17). Two i.v. does of 16 to 160 pmol/hr (16 and 160 times greater than the icv dose) were tested and both attenuated neuronal cell death significantl ⁇ ( ⁇ 0.01 ).
  • neuronal cell death in the hippocampus resulting from forebrain ischemia/reperfusion in rats was significantl ⁇ attenuated b ⁇ iv infusion of PACAP38.
  • Administration beginning 24 hrs after the ischemia/reperfusion was as effective as infusion starting immediatel ⁇ after the ischemia/reperfusion.
  • ADDRESSEE DAVID G. CONLIN; DIKE, BRONSTEIN,
  • MOLECULE TYPE peptide

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Abstract

L'invention porte sur un procédé et des préparations pharmaceutiques pour le traitement et la prévention de lésions des neurones chez les mammifères, ledit procédé consistant à administrer une dose efficace de PACAP ou de l'un de ses agonistes, analogues ou dérivés présentant l'activité neurotrophique du PACAP, placé dans un excipient pharmacocompatible, en une concentration efficace pour protéger in vivo les neurones.
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WO1997040388A1 (fr) * 1996-04-25 1997-10-30 Shiseido Company, Ltd. Peptides presentant une affinite specifique pour les recepteurs pacap de type 1
WO1998032857A1 (fr) * 1997-01-23 1998-07-30 University Of Victoria Innovation And Development Corporation Gene de neuropeptides de poulet servant a la production de volaille
WO2000005260A1 (fr) * 1998-07-20 2000-02-03 Societe De Conseils De Recherches Et D'applications Scientifiques Sas Analogues peptidiques des pacap
US6242563B1 (en) 1998-07-20 2001-06-05 Societe De Conseils De Recherches Et D'applications Scientifiques, Sas Peptide analogues
EP1752158A4 (fr) * 2004-04-23 2009-08-05 Senju Pharma Co Promoteur de la neuritogénèse de la cornée contenant du pacap et son dérivé
WO2009046876A3 (fr) * 2007-09-11 2009-11-05 Mondobiotech Laboratories Ag Utilisation d'un peptide en tant qu'agent thérapeutique
WO2009033767A3 (fr) * 2007-09-11 2009-11-12 Mondobiotech Laboratories Ag Utilisation d'un peptide en tant qu'agent thérapeutique
WO2011054001A2 (fr) 2009-11-02 2011-05-05 The Administrators Of The Tulane Analogues de polypeptide d'activation d'adénylate cyclase hypophysaire (pacap) et procédés pour leur utilisation
US9006181B2 (en) 2004-07-21 2015-04-14 The Administrators Of The Tulane Educational Fund Treatment of renal dysfunction and multiple myeloma using PACAP compounds
CN114173874A (zh) * 2019-05-14 2022-03-11 千寿制药株式会社 含pacap肽或pacap的稳定化肽的神经营养性角膜炎的预防或治疗用组合物

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AU2002325712C1 (en) 2001-08-30 2008-07-31 Stem Cell Therapeutics Inc. Differentiation of neural stem cells and therapeutic use theeof
JP4906231B2 (ja) 2001-09-14 2012-03-28 ステム セル セラピューティクス インコーポレイテッド プロラクチン誘導性の神経幹細胞数の増加ならびにその治療用途
AU2003228050A1 (en) * 2002-05-03 2003-11-17 Neuronova Ab Therapeutic use of PACAP, Maxadilan, PACAP receptor agonist and/or ADCYAP1R1 in the treatment of CNS disorders
EP1546198A1 (fr) * 2002-07-31 2005-06-29 Stem Cell Therapeutics Inc. Procede d'amelioration de la proliferation, de la differentiation et de la survie de cellules souches neurales au moyen du polypeptide d'activation d'adenylate cyclase hypophysaire (pacap)
WO2005077404A1 (fr) 2004-02-13 2005-08-25 Stem Cell Therapeutics Corp. Utilisation de l'hormone de luteinisation (lh) et de la gonadotropine chorionique (hcg) pour la proliferation de cellules souches neuronales et la neurogenese
JP2009509943A (ja) 2005-09-27 2009-03-12 ステム セル セラピューティクス コーポレイション プロラクチンにより制御される乏突起膠細胞前駆体細胞の増殖
EP2004211A4 (fr) 2006-03-17 2010-07-07 Stem Cell Therapeutics Corp Régimes de dosage de lh ou hcg et epo pour le traitement de troubles neurologiques
KR20100059867A (ko) * 2007-09-11 2010-06-04 몬도바이오테크 래보래토리즈 아게 치료제로서의 밴드 ３ 단백질 및 ｐａｃａｐ－２７의 용도
ITMI20110583A1 (it) 2011-04-08 2012-10-09 Hmfra Hungary Ltd Liability Company Preparazioni oftalmiche a base di pacap (pituitary adenylate cyclase activating polypeptide) al fine di ripristinare la normale funzione visiva nel glaucoma in fase precoce
JP7695772B2 (ja) * 2017-11-14 2025-06-19 千寿製薬株式会社 Ｐａｃａｐの安定化ペプチド

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Cited By (15)

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Publication number Priority date Publication date Assignee Title
WO1997040388A1 (fr) * 1996-04-25 1997-10-30 Shiseido Company, Ltd. Peptides presentant une affinite specifique pour les recepteurs pacap de type 1
US6165755A (en) * 1997-01-23 2000-12-26 University Of Victoria Innovation And Development Corporation Chicken neuropeptide gene useful for improved poultry production
WO1998032857A1 (fr) * 1997-01-23 1998-07-30 University Of Victoria Innovation And Development Corporation Gene de neuropeptides de poulet servant a la production de volaille
EP2161282A1 (fr) * 1998-07-20 2010-03-10 Ipsen Pharma Peptides analogues de PACAP
US6242563B1 (en) 1998-07-20 2001-06-05 Societe De Conseils De Recherches Et D'applications Scientifiques, Sas Peptide analogues
JP2002521390A (ja) * 1998-07-20 2002-07-16 ソシエテ・ドゥ・コンセイユ・ドゥ・ルシェルシュ・エ・ダプリカーション・シャンティフィック・エス・ア・エス Ｐａｃａｐのペプチド類似体
WO2000005260A1 (fr) * 1998-07-20 2000-02-03 Societe De Conseils De Recherches Et D'applications Scientifiques Sas Analogues peptidiques des pacap
EP1752158A4 (fr) * 2004-04-23 2009-08-05 Senju Pharma Co Promoteur de la neuritogénèse de la cornée contenant du pacap et son dérivé
US9006181B2 (en) 2004-07-21 2015-04-14 The Administrators Of The Tulane Educational Fund Treatment of renal dysfunction and multiple myeloma using PACAP compounds
WO2009046876A3 (fr) * 2007-09-11 2009-11-05 Mondobiotech Laboratories Ag Utilisation d'un peptide en tant qu'agent thérapeutique
WO2009033767A3 (fr) * 2007-09-11 2009-11-12 Mondobiotech Laboratories Ag Utilisation d'un peptide en tant qu'agent thérapeutique
WO2011054001A2 (fr) 2009-11-02 2011-05-05 The Administrators Of The Tulane Analogues de polypeptide d'activation d'adénylate cyclase hypophysaire (pacap) et procédés pour leur utilisation
EP2496245A4 (fr) * 2009-11-02 2013-08-21 Univ Tulane Analogues de polypeptide d'activation d'adénylate cyclase hypophysaire (pacap) et procédés pour leur utilisation
US8916517B2 (en) 2009-11-02 2014-12-23 The Administrators Of The Tulane Educational Fund Analogs of pituitary adenylate cyclase-activating polypeptide (PACAP) and methods for their use
CN114173874A (zh) * 2019-05-14 2022-03-11 千寿制药株式会社 含pacap肽或pacap的稳定化肽的神经营养性角膜炎的预防或治疗用组合物

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