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WO1997049422A1 - Procede d'utilisation de ligands du recepteur opioïde kappa - Google Patents

Procede d'utilisation de ligands du recepteur opioïde kappa Download PDF

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Publication number
WO1997049422A1
WO1997049422A1 PCT/US1997/010916 US9710916W WO9749422A1 WO 1997049422 A1 WO1997049422 A1 WO 1997049422A1 US 9710916 W US9710916 W US 9710916W WO 9749422 A1 WO9749422 A1 WO 9749422A1
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dyn
dynoφhin
blood
opioid receptor
kappa opioid
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PCT/US1997/010916
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English (en)
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Mary Jeanne Kreek
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The Rockefeller University
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Priority to AU34999/97A priority Critical patent/AU3499997A/en
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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
    • A61K38/33—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans derived from pro-opiomelanocortin, pro-enkephalin or pro-dynorphin

Definitions

  • Dyn A (1-13) in humans have shown that this peptide caused elevation in serum levels of prolactin. which reflects diminished dopaminergic tone in hypothalamus (Kreek et al . "Dynorphin A, n administration causes elevation ot serum levels of prolactin in human subjects In Problems of Drug Dependence 1993 Proceedings of the 55th Annual Scientific Meeting. The College on Problems of Drug Dependence. Inc . NIDA Research Monograph
  • Drugs of abuse such as cocaine are known to increase extracellular fluid dopamine levels in st ⁇ atum and nucleus accumbens of rats (Maisonneuve and Kreek, "Acute tolerance to the dopamine response induced by a binge pattern of cocaine administration in male rats an in vivo microdialysis study " J Pharmacol Exp Ther. 268: 916-921. 1994).
  • Cocaine also alters the dynorphin systems in rats: preprodynorphin mRNA is increased in the caudate putamen (Spangler et al. , " 'Binge' cocaine administration induces a sustained increase of prodynorphin mRNA in rat caudate-putamen" Mol. Brain Res.
  • kappa opioid receptor mRNA is decreased in the substantia nigra (Sprangler et al. , "Kappa opioid receptor mRNA levels in the rat brain: effects of dopamine antagonists and cocaine" In E.J. Simon (Ed.), Regulatory Peptides. Vol. 54, Elsevier, Amsterdam, pp. 283, 1994 and Spangler et al , "Regulation of kappa opioid receptor mRNA in the rat brain by "binge” pattern cocaine administration and correlation with preprodynorphin mRNA” Mol. Brain Res. , 1996, 38:71-76.
  • the present invention relates to the treatment and management of diseases states which are resultant from the acute and chronic abuse and addiction to cocaine and 97 / 49422 PC17US97/10916
  • this invention relates to a method of reducing basal dopamine levels or attenuating surges of dopamine levels in mammalian brain which comprises administering to a patient in need of such therapy a pharmaceutically acceptable amount of a compound which is a ligand of the kappa opioid receptor.
  • the method of the instant invention involves the treatment of patients who exhibit enhanced levels of dopamine or surges in dopamine levels in their brains. Typical of such patients are acute and chronic abusers, and addicts of cocaine. Other stimulants, such as the amphetamine-type stimulants, have similar effects upon the normal levels of dopamine in the brains.
  • Certain patients having behavioral disorders exhibit similar patterns of enhanced dopamine levels or surges in dopamine levels in the brain. Accordingly, the method of the present invention likewise finds utility in the treatment of such patients, especially in those who abuse, or are addicted to, nicotine, marijuana and alcohol.
  • Other behavioral disorders such as excess consumption of food, opioid abuse and addiction, and compulsive gambling, which exhibit similar variation in normal dopamine levels, likewise can be treated by the methods of the instant invention.
  • the present invention comprises a method of reducing basal dopamine levels or attenuating surges of dopamine levels in mammalian brain which comprises administering to a patient in need of such therapy a pharmaceutically acceptable amount of a compound which is a ligand of the kappa opioid receptor.
  • the ligands of the kappa opioid receptor useful in the practice of the instant invention can be selected from the group consisting of known ligands of the kappa opioid receptor, such as dyno ⁇ hin A, active fragments thereof, and synthetic analogs thereof.
  • dyno ⁇ hin A, (dynA) having the structure
  • Dyno ⁇ hin A as well as the Dyno ⁇ hin A(l-6), Dyno ⁇ hin A(l-7).
  • Dyno ⁇ hin A(l-8), Dyno ⁇ hin A(l-9), Dyno ⁇ hin A(l-10), and Dyno ⁇ hin A(l-13) peptides are known in the art and are commercially available from, for instance. ICN Biomedical, Inc.
  • dyno ⁇ hin A analogs which can be utilized in the practice of the instant invention are the synthetic analogs of Dyno ⁇ hin A which contain various substituted amino acids in place of the naturally occurring amino acid, a different amino acid in place of an individual amino acid, and/or a D-amino acid substituted for an individual amino acid.
  • these analogs can be of the full seventeen amino acid sequence, or can be only a portion thereof.
  • a particularly preferred dyno ⁇ hin analog which has found utility in the present invention is the dyno ⁇ hin A analog which is fN-methyl-Tyr' ,N-methyl-Arg 7 -D- Leu 8 JDynA (1-8) ethylamide having the sequence
  • This compound finds particular utility in the instant invention due to its ability to cross the blood- brain barrier, and its resistance to degradation by mammalian enzymes, thus making it longer lived when administered in vivo.
  • the peptides useful in this invention can be, and have been, synthesized by known solid phase techniques. See, for example, Barany and Merrifield (1979) in The Peptides, eds. Gross and Meienhofer (Academic Press, New York) Vol. 2A,
  • the products can be prepared by manual methods or, for example, on a peptide synthesizer such as the Applied Biosystems 430 unit. Preparation of certain of the dynorphin analogs is described in the aforementioned U. S. Patents 4,707,468, 4,551 ,273 and 4,481 ,138.
  • the ability of the kappa opioid ligands of the instant invention to reduce basal dopamine levels or attenuate surges of dopamine levels in mammalian brain and thereby modulate the effects of cocaine dependency can be determined by an assay which evaluates the effects of cocaine on ppDyn mRNA, in the presence or absence of the exogenously administered dyno ⁇ hin A, a fragment thereof, or a synthetically derived analog thereof.
  • assays can thus be utilized as a method of screening for agents which can be utilized for the treatment of acute or chronic abuse and/or addiction to stimulants, or a behavioral disorder, which results in enhanced dopamine levels or surges in dopamine levels in the brain of a patient.
  • the products of this invention will generally be administered in the same manner as other bioactive peptides, i.e. , parenterally. intraventricularly, intrathecally, or by infusion. Since their chemical structure is similar to other bioactive peptides. they will generally be administered with the same types of pharmaceutically acceptable excipients.
  • the peptides useful in the methods of the present invention are amphoteric, they may be utilized as free bases, as acid addition salts, or as metal salts.
  • the salts must, of course, be pharmaceutically acceptable, and these will include metal salts, particularly alkali and alkaline earth metal salts, and typically potassium or sodium salts.
  • metal salts particularly alkali and alkaline earth metal salts, and typically potassium or sodium salts.
  • a wide variety of pharmaceutically acceptable acid addition salts are available. These include those prepared from both organic and inorganic acids, preferably mineral acids. Typical acids, which may be mentioned by way of example, include citric, succinic, lactic, hydrochloric and hydrobromic acids. Such products are readily prepared by procedures well known to those skilled in the art.
  • the ligands of the kappa opioid receptor useful in the methods of the invention will normally be provided for as parenteral compositions for injection or infusion. They can, for example, be suspended in an inert oil, suitably a vegetable oil such as sesame, peanut, or olive oil. Alternatively, they can be suspended in an aqueous isotonic buffer solution at a pH of about 5.6 to 7.4. Useful buffers include sodium citrate-citric acid and sodium phosphate-phosphoric acid.
  • the desired isotonicity may be accomplished using sodium chloride or other pharmaceutically acceptable agents, such as dextrose, boric acid, sodium tartrate, propylene glycol, or other inorganic or organic solutes.
  • sodium chloride is preferred, particularly when the buffer contains sodium ions.
  • the solutions may be thickened with a thickening agent such as methyl cellulose.
  • a thickening agent such as methyl cellulose.
  • They may be prepared in emulsified form, either water in oil or oil in water. Any of a wide variety of pharmaceutically acceptable emulsifying agents may be employed including, for example, acacia powder, or an alkaryl polyether alcohol sulfate or sulfonate such as Triton.
  • delayed and sustained release formulations which can provide administration of the desired active agent over a period of several hours, thus obviating the need for multiple doses, and provided a sustained level of the active over the therapeutic period.
  • compositions of the invention are prepared by mixing the ingredients following generally accepted procedures.
  • the selected components may be simply mixed in a blender or other standard device to produce a concentrated mixture which may then be adjusted to the final concentration and viscosity by the addition of water or thickening agent and, optionally, a buffer to control pH or an additional solute to control tonicity.
  • compositions will be provided in unit dosage form containing an amount of the ligand of the kappa opioid receptor which will be effective in one or multiple doses to reduce basal dopamine levels or attenuate surges of dopamine levels in mammalian brain.
  • an effective amount of the therapeutic agent will vary with many factors, including the age and weight of the patient, the dopamine levels already present, the potency of the selected dyno ⁇ hin A. fragment or analog thereof, and other factors.
  • Typical dosage units will contain from 0.2 to 0.8 mg/ml, although wide variations from this range are possible while yet achieving useful results.
  • Dyn A (7-17) Dyn A (8-17), Dyn A (6-17) and Dyn A (1-7).
  • Dyn A (2-17) was found to be the major non- opioid biotransformation product.
  • Dyn A (2-17) was further biotransformed into Dyn A (3-17) and Dyn A (4-17). However, this latter conversion occurred very slowly.
  • Another major biotransformation pathway is the cleavage of the peptide linkage between Arg(6) and Arg(7) to produce the opioid peptide Dyn A (1-6) and the non-opioid peptide Dyn A (7-17). Both of these biotransformation products were further processed into minor products Dyn A (1-6) to Dyn A (2-6); and Dyn A (7-17) to Dyn A (8-17), Dyn A (9-17) and Dyn A (11-17).
  • Two types of enzymes are thought to be mainly responsible for Dyn A peptide biotransformation: amino ⁇ eptidebase(s) (Benuck et al.
  • Dyn A (1-17) in vitro in rhesus monkey blood and in human blood was found to be very similar.
  • Two major biotransformation pathways were identified for Dyn A (1-17) in vitro processing in rhesus monkey blood: cleavage of N-terminal Tyr(l) to form Dyn A (2-17), and cleavage between Arg(6) and Arg(7) to form Dyn A (1-6) and Dyn A (7-17).
  • Dyn A (2-17) was the major biotransformation product both in human and rhesus monkey blood. Although it does not bind to opioid receptors (Walker et al. , "Nonopiate effects of dyno ⁇ hin and des-Tyr-dynorphin" Science, 218: 1136-1 138, 1982), Dyn A (2-17) has been shown to produce a profile of pharmacological activities, capable of potent physiological effects, in part similar to those of the full-length opioid peptide, Dyn A (1-17). Dyn A (2-17) was shown to stimulate many of the pathophysiological consequences of Dyn A (1 -17) such as motor and electrophysiological effects when administered to rats (Walker et al. , 1982).
  • Dyn A (2-17) administered through a microdialysis probe placed in rat hippocampus caused a dose-related increase in the extracellular levels of excitatory amino acids, glutamate and aspartate (Faden, "Dyno ⁇ hin increases extracellular levels of excitatory amino acids in the brain through a non-opioid mechanism" J. Neurosci. 12: 425-429, 1992).
  • Dyn A (2-17) was shown to be as effective as the active opioid Dyn A (1-17) to suppress naloxone-precipitated withdrawal jumping in morphine-dependent mice (Takemori et al. , 1993).
  • Dyn A (2-17) Comparative studies of Dyn A (2-17) with the opioid-active but shortened form Dyn A (1-13) indicated that both peptides were effective in inhibiting the expression of physical dependence on morphine in mice after acute, high-dose exposure and chronic exposure. Both Dyn A (1-13) and Dyn A (2-17) effectively suppressed the expression of chronic opiate tolerance (Hooke et al. , 1995a). Dyn A (2-17) was shown to have antinoicceptive effect in the writhing assay, similar to Dyn A (1-13) when injected to mice by the intravenous, intrathecal, intracerebro ventricular and intraperitoneal routes (Hooke et al. , "[Des-Tyr 1 ]
  • Dyno ⁇ hin A-(2-17) has naloxone-insensitive antinociceptive effect in the writhing assay. " /. Pharmacol. Exp. Therap. 273: 802-807, 1995b). Studies have indicated that the behavioral effects of Dyn A (1-13) were generally unaffected by naloxone (Walker et al. , "Behavioral effects of dyno ⁇ hin 1 u in the mouse and rat: initial observations" Peptides, 1: 341-345, 1980). It was suggested that within Dyn A (1-17). Two biological active sequences existed, on opiate and the other non-opiate, both of which were able to produce significant behavioral activity (Walker et al. , 1982).
  • Dyn A (1-17) was much more rapidly transformed in rhesus monkey blood than in human blood.
  • Dyn A (1-17) gradually disappeared with incubation time, but still could be detected in the blood of some subjects at 240 minutes incubation. By comparison, Dyn A (1-17) rapidly disappeared in rhesus monkey blood.
  • Dyn A (2-17) the major non-opioid biotransformation product, was the long-lasting Dyn A fragment both in human and rhesus monkey blood. Compared to the biotransformation kinetics in rhesus monkey blood, Dyn A (2-17) formed more slowly in human blood but lasted much longer. It still could be detected in human blood at 240 minutes incubation. The observation that different species gave rise to differences in Dyn A processing was previously reported in studies of degradation of Dyn A (1-17) in vitro in rat and guinea pig brain tissue.
  • Dyn A (1-6) and Dyn A (7-17) were formed during the incubation of Dyn A (1-17), but rapidly disappeared.
  • Dyn A (1-6) and Dyn A (7-17) disappeared within 180 minutes of incubation in human blood and 45 minutes in rhesus monkey blood.
  • a strong correspondence was observed between the kinetics of appearance and disappearance of Dyn A (1-6) and that of dyn A (7-17).
  • Dyn A (7-17) was relatively slow at 25 °C, suggesting that the enzyme(s) responsible for Arg(6)-Arg(7) cleavage had relatively low activity at this incubation temperature.
  • Dyn A (7-17) was formed with an hour of incubation, and rapidly disappeared.
  • a temperature dependence in Dyn A (1-13) biotransformation by membrane-bound rat brain enzyme had previously been reported (Leslie and Goldstein, 1982). The authors found that the rate of metabolism was reduced by incubation at 0°C.
  • Rhesus monkeys as a primate model have been used in many behavioral studies of the effects of Dyn A ( 1-13).
  • the effects of Dyn A (1-13) were studied in mo ⁇ hine-dependent rhesus monkeys (Aceto et al. , "Dyno ⁇ hin-(l-13): Effects in nontolerant and morphine-dependent rhesus monkeys", Eur. J. Pharmacol. 83: 139-142, 1982). These studies indicated that Dyn A (1-13) suppressed withdrawal signs in a dose-related manner within 30 minutes. In non-mo ⁇ hine-dependeru monkeys, Dyn A (1-13) did not produce typical mo ⁇ hine effects. Studies also showed that Dyn A (1-13) effects were significantly decreasing at 90 minutes .
  • Dyn A peptides both the shortened Dyn A (1-13) and the natural full length Dyn A (1 -17), can be biotransformed in a biological system into both opioid and also non-opioid Dyn A fragments, some of which have opioid-like activities (Chou et al. , "Study of opioid peptides by laser deso ⁇ tion mass spectrometry" In: Problems of Drug Dependence 1992: Proceedings of the 54th Annual Scientific Meeting, The College on Problems of Drug Dependence, Inc. , NIDA Research Monograph Series, Vol.
  • Blood was obtained from seven healthy volunteers, five men and two women with an average age of 32 years (range 25- 47). Blood was obtained from three healthy adult female rhesus monkeys, average age 8.3 years (range 7-10).
  • Dyn A (1-17) was synthesized by Chiron Mimotopes Peptides Systems (San Diego. CA), and made available by the National Institute on Drug Abuse (Rockville, MD).
  • Dyn A (2-17) was synthesized by American Peptide Company (Sunnyvale, CA), and made available by Neurobiological Technologies Inc. (Richmond. CA).
  • Dyn A (1-12) was purchased from Peninsula Laboratories, Inc. and made available by Neurobiological Technologies, Inc.
  • Somatostatin was purchased from Sigma Chemical Company (St. Louis, MO). Saline (0.9% NaCl) was from Abbott Laboratories (North Chicago, IL).
  • ⁇ -Cyano-4-hydroxycinnamic acid (4HCCA) was obtained from Aldrich Chemical Company, Inc. (Milwaukee, WI).
  • HPLC-grade acetonitrile was purchased from Burdick & Jackson
  • Dynorphin A processing and subsequent sample preparation. Blood was drawn from subjects into vacutainer tubes preconditioned with EDTA (Becton Dickisnon. Rutherford, NJ). Typically, an aliquot 12.0 ml of freshly drawn blood was transferred to a 50 ml Falcon polypropylene tube (Becton Dickinson, Lincoln Park, NJ). Following removal of 1.0 ml blood for a control, 7.08 mg of Dyn A (1-17), dissolved in 500 ⁇ lj of saline, was immediately added to the remaining blood (i.e. 0.59 mg/ml, or 196.4 ⁇ M).
  • the internal standard was used as the calibration reference for the sample filtration process and subsequent mass and subsequent mass spectrometric analysis. However, the use of this internal standard would not allow for determination of whether or not Dyn A (1-17) or any biotransformation products were bound to blood cells and thus lost in the centrifugation step.
  • Mass spectrometry Biotransformation products were identified and the kinetics of the biotransformation processes elucidated by a mass spectrometer consisting of a matrix-assisted laser deso ⁇ tion ion source, coupled with a linear time-of-light mass analyzer (MALDI-MS), details of which have been described elsewhere (Beavis et al. , Rapid Commun. Mass Spectrom. 3 :233-237 ' , 1989, and Beavis, et al. . Anal. Chem. 62: 1836-1840, 1990).
  • plasma samples with acetonitrile (2: 1) were mixed with matrix solution.
  • Matrix solution was made by dissolving excess amount (5 mg/ml) of 4HCCA in acetonitrile and 0.1 % aqueous TFA [1 :2 (v/v)] to make a saturated solution.
  • Sample was mixed with matrix solution in a ration of 1 :2 (v/v).
  • a 0.5 ⁇ l aliquot of sample-matrix solution was applied to the mass spectrometer sample probe tip, and allowed to evaporate to dryness in the air. (In the current instrument, the mass spectrometer sample probe tip allows for the simultaneous loading of 10 separate sample spots, each one 2 mm in diameter - allowing for rapid sequential determination of multiple samples).
  • the sample probe was inserted into the mass spectrometer vacuum system where solvents in the sample were completely removed. After 5 minutes, a working pressure of approximately 10 7 torr was achieved. To obtain adequate statistics, the results from 200 laser shots were added for each mass spectrum. In this study, on spectrum was obtained for each sample from each subject.
  • the MALDI-MS system was calibrated with a series of Dyn A peptides: Dyn A (1-17), Dyn A (2-17), Dyn A (1-12) and Dyn A ( 1-16). Serial dilution was performed to make fourteen different concentrations of the peptide standards. A constant amount of internal standard (0.516 mg/ml, or 7.88 nmol somatostain) dissolved in 25 ⁇ l of 0.05 % TFA was added to 500 ⁇ l of each concentration of standard solutions (i.e. 24.6 ⁇ g/ml somatostain). Mass spectrometric measurements were carried out in triplicate for each standard solution. The ratio of the peak height of each Dyn A standard to the peak height of the internal standard and the standard error of the triplicate measurements were calculated.
  • Biotransformation kinetics of Dyn A peptides The kinetics of Dyn A biotransformation in vitro in human and rhesus monkey blood was studied with MALDI-MS.
  • Dyn A (1-17) was added to the freshly drawn blood to yield a concentration of 0.59 mg/ml. The blood was incubated at 37 °C. Aliquots were withdrawn before and at specific time points after the addition of Dyn A (1-17). Mass spectrometric measurements were carried out with the internal standard. 31 .5 nmol somatostatin. being added to 2.0 ml of the sample solution (i.e. 24.6 ⁇ g/ml).
  • the relative peak height of a dyno ⁇ hin A fragment to the peak height of the internal standard was used as a measure of the abundance of that fragment.
  • the values of relative peak height of Dyn A (1-17) and its fragments, Dyn A (2-17) Dyn A (1-6) and Dyn A (7-17), were plotted versus the incubation time.
  • the curve fitting was constructed with nonlinear curve-fitting routine using KaleidaGraphTM (Version 3.0.5, Abelbeck Software, Reading, PA).
  • Dyn A (1-17) in vitro biotransformation in human blood was biotransformed into a variety of dynorphin A fragments upon incubation at 37 °C in human blood.
  • Table 1 summarizes the biotransformation products of Dyn A (1- 17) detected in seven human subjects. The major biotransformation products were identified to be Dyn A (2-17), Dyn A (1 -6) and Dyn A (7-17). These products were further processed into minor products: Dyn A (2-17) to Dyn A(3-17) and Dyn A (4-17); Dyn A (1-6) to Dyn A (2-6); Dyn A (7-17) to Dyn A (8-17), Dyn A (9-17) an Dyn (1 1-17). Other minor biotransformation products, such as Dyn A (8-15), Dyn A (9-15), Dyn A (1-11), Dyn A (1 -12) and Dyn A (4-1 1), were also detected in blood of some human subjects.
  • EXAMPLE 2 The present study was planned to reevaluate the effects of cocaine on ppDyn mRNA and to use the recently cloned rat KOR cDNA, to study the effects of cocaine administration on KOR mRNA levels in the caudate-putamen, the substantia nigra and the ventral tegmental area. These studies were performed with a quantitative mRNA assay utilizing trichloroacetic acid precipitation of RNase-protected cRNA:mRNA hybrids. This assay allows analysis of multiple genes in a single region and facilitates correlation analysis between mRNAs on an individual animal basis both within and between brain regions.
  • mice Male Fischer rats (60 days old; Charles River, NY) were adapted for 7 days to a 12 h light/dark cycle in a stress minimized facility with handling once a day in the morning. To control for possible injection effects, all rats received an equal number of injections during 14 days. Rats received either saline (1 ml/kg. day) or cocaine (45 mg/kg. day) in three daily administrations 1 hour apart (9:30, 10:30 and 11 :30 a.m.) starting 30 minutes after the lights were turned on.
  • RNA concentrations were determined by hybridization of duplicate diluted extracts to a labeled anti-sense cRNA probe complementary to human 18S rRNA as described previously [2], using plasmid pS/E provided by Drs. Timothy Nilsen and Patricia Maroney of Case Western Reserve University.
  • This plasmid contains the Sall-EcoRl fragment spanning most of the 18S rna gene inserted downstream O 97/49422 PC17US97/10916
  • the 18S ma is among the slowest evolving sequences found throughout living organisms
  • the human 18S ⁇ boprobe hybridizes very effectively to lat 18S ina, showing a single band on an acrylamide/urea gel following RNase tieatment of cRNA rna hybrids (not shown)
  • both methods produced similar values, with r > 0 9
  • the 18S calibration cuive was linear from 2 5 to 60 ng Aliquots ol expenmental samples were diluted to approximately 2 ng/ ⁇ l and 4 ⁇ l weie assayed in duplicate
  • the standard curve samples for establishing the linear regression values extended from 80 to 0.625 pg/aliquot by two fold dilutions. Duplicates of these standards were included with every assay set, with a minimum of four zero tubes. All calibration samples included 10 ⁇ g of E. coli tRNA. Input probe was always present at 10-fold excess concentration above the highest standard and greater than 20-fold excess above the level of the highest sample tested, to assure that the samples were in the linear range of the calibration curve.
  • SAL14 One group of rats (SAL14) served as control for two cocaine treated groups (SAL11-COC3 and COC14) in the first experiment described. All of these animals received 42 injections of saline and/or cocaine. As we had determined that corticosterone levels were elevated for up to one week by saline injections, the SAL11 -COC3 animals were pretreated with saline for 11 days in order to desensitize the animals to the effects of injections. For this reason, both cocaine groups were treated in a way that allowed them to be compared to the single control group that had been treated with saline for 14 days. Therefore, statistical calculations were made comparing the two cocaine treated groups to the one saline treated group; i.e.
  • the 11-day aline-3 day cocaine group together with the 14-day cocaine group vs. the 14-day saline group (SAL1 1 -COC3 and COC14 vs. SAL14).
  • Analyses of variance followed by planned orthogonal comparisons were used to determine whether the mean differences were significant between mRNA levels of specific brain regions between the treatment groups.
  • Least squares correlation analysis was carried out to determine Pearson's coefficient for levels of paired mRNAs of genes of interest in a given brain region (ppDyn X KOR in caudateputamen ) . or in different brain regions (ppDyn in caudate-putamen X Kor in substantia nisra).
  • Table 3 shows the levels of KOR mRNA in two midbrain regions of the same animals shown in Table 2.
  • KOR mRNA levels in the adjoining ventral tegmental area were not significantly altered by cocaine administration.
  • cocaine does not have a generalized effect on KOR mRNA.
  • Densities are presented as attomole mRNA/ ⁇ g total RNA + SEM.
  • Table 3 shows the results of a study carried out to determine the persistence of the ppDyn and KOR alterations found in response to 'binge' cocaine for 14 days and then kept for 10 days without further injections or handling. There were no significant changes in ppDyn or ppEnk mRNA in the caudate-putamen. or KOR mRNA levels in the substantia nigra, in the groups which had 10 days of no treatment following 14 days of 'binge' cocaine injections (SAL14-W10 vs. COC14-W10). In addition, levels of these three genes were not significantly different from the levels seen in the saline-injected animals from the first protocol (compare Tables 1 and 2 with Table 3). These results indicate that the significant modulations in ppDyn in the caudate-putamen and KOR in the substantia nigra seen in rats treated with cocaine for 14 days do not persist 10 days after cessation of cocaine administered.
  • KOR agonists including dyno ⁇ hin A 1-13 and 1-17, reduce dopamine release caused by activators.
  • You et al. recently reported that a Dl dopamine receptor agonist administered into the forebrain (the striatum) led to an increase in extracellular dyno ⁇ hin B in the midbrain (the substantia nigra).
  • the same group earlier reported that intranigral injections of dyno ⁇ hin A decreased striatal dopamine levels between 40 and 80 minutes after injection.
  • the firing rate of midbrain dopamine neurons in the ventral tegmental area and substantia nigra is lowered within minutes of i.v. cocaine administration.
  • Dendritic release of dopamine in the midbrain could lower dopaminergic cell activity by binding to Dl type dopamine receptors acting presynaptically at GABAergie terminals and D2 type somatodendritic autoreceptors on dopaminergic cells.
  • Dl dopamine receptor agonists facilitate GABA release in the midbrain, an action that hype ⁇ olarizes postsynaptic dopaminergic cells.
  • Evidence for autoreceptor inhibition of dopaminergic cell activity was provided by use of D2 dopamine receptor antagonists, which cause increased firing of dopaminergic cells , reportedly by blocking somatodendritic autoreceptors in the midbrain.
  • Hype ⁇ olarization of dopaminergic cells by GABA acting at GABA B receptors and by dopamine acting at D2 autorecptors has been proposed to be effected by a common mechanism. It has been suggested that the binding of dopamine in the midbrain to Dl presynaptic dopamine receptors (facilitating GABA release) and to somatodendritic D2 autoreceptors can act synergistically to lower dopaminergic transmission.
  • KOR levels have been decreased (for example, by chronic "binge" cocaine administration, as shown in the substantia nigra in the present study), there would be less inhibition of dopamine release and a consequent higher extracellular level of dopamine in the substantia nigra.
  • a recent study reported by Kalivas and Duffy indicates that there is an increased midbrain dopamine level following chronic cocaine administration.
  • KOR in the substantia nigra would result in: (1) increased extracellular dopamine in the substantia nigra; (2) increased binding to dopamine receptors downmodulating dopaminergic cell activity in the substantia nigra; (3) decreased firing of dopaminergic cells; and (4) decreased dopamine release at the nigrostriatal terminals.
  • This model is in accord with recent data demonstrating a reduced baseline of extracellular dopamine in the striatum following 14-day "binge" cocaine administration.
  • EXAMPLE 3 The objective of this study was to characterize the potential role of the endogenous peptide Dyno ⁇ hin A M7 (Dyn A uv ) in modulating dopaminergic neurotransmission in the nucleus accumbens (NAcc).
  • Male Fischer 344 rats were maintained on a 12 hour light; dark cycle with food and water ad libitum. Rats were implanted stereotaxically with one guide cannula above the Nacc. A calibrated microdialysis probe was inserted through the guide cannula of each animal the night before dialysis experiment. Artificial cerebrospinal fluid (aCSF) was delivered by pump at a constant rate of 1 ul/min. On the day of the experiment, six 20 minute baseline samples were taken.
  • aCSF artificial cerebrospinal fluid
  • EXAMPLE 4 The objective of this study was to characterize the potential role of the endogenous peptide Dyno ⁇ hin A,. 17 (Dyn Aj. ⁇ ) in modulating dopaminergic neurotransmission in the nucleus accumbens (NAcc).
  • NAcc nucleus accumbens
  • Male Fischer 344 rats were deemed for at least four days to controlled conditions. Animals were maintained on a 12 hour light:dark cycle with food and water provided ad libitum. Rats were implanted stereotaxically with one guide cannula above the NAcc. A calibrated microdialysis probe was inserted through the guide cannula of each animal the night before the dialysis experiment.
  • aCSF Artificial cerebrospinal fluid
  • Dyn A dyno ⁇ hin A
  • 4HCCA ⁇ -cyano-4-hydroxycinnamic acid
  • Leu leucine
  • Phe phenylalanine
  • Gly glycine
  • MALDI-MS matrix-assisted laser deso ⁇ tion/ionization mass spectrometry.
  • Dyn A (1-8) ethylamide (E-2078) was synthesized and kindly supplied by Eisai Co. Ltd. (Ibaraki, Japan).
  • Dyn A (1-8) was purchased from Peninsula Laboratories, Inc. (Belmont, CA).
  • Angiotensin III and diphenhydramine HCl were purchased from Sigma Chemical Co. (St. Louis, MO).
  • Ketamine HCl was obtained from Fort Dodge (Fort Dodge, IA). Heparin was purchased from Elkins- Sinn (Cherry Hill, NJ). Saline (0.9% NaCl) and dextrose were from Abbott Laboratories (North Chicago, IL).
  • 4HCCA was obtained from Aldrich Chemical company, Inc. (Milwaukee, WI). High-performance liquid chromatography grade acetonitrile was purchased from Burdick & Jackson (Muskegon, MI) and TFA from Fisher Scientific (Fair Lawn, NJ).
  • plasma was separated from the blood by centrifugation in a Sorvall RC-5B refrigerated (0-5 °C) superspeed centrifuge (Sorvall Instruments, Du Pont Company, Newtown, CT), at 3,400 x g for 5 minutes. A total of 200 ⁇ l of plasma containing E-2078 and proteolytic products was added to 1.8 ml of 1.0% TFA aqueous solution. The samples were frozen at -70°C until analysis.
  • E-2078 In Vivo Studies in Rhesus Monkeys. Monkeys received injections with diphenhydramine (1.2 mg/kg, intramuscularly) as a pretreatment to limit the possible consequences of histamine release after administration of relatively large amounts of E-2078. Thirty minutes later, they were anesthetized with ketamine (10 mg/kg, i.m.). The back of the lower leg area was carefully shaved to avoid any adventitious bleeding.
  • An indwelling catheter (Angiocath, 22 gauge, 1-inch long, Becton Dickinson, Sandy, UT) was acutely placed in a superficial veining of each leg, secured and flushed with heparinized saline (20 U/ml), and connected to a multisample injection port.
  • One milliliter of blood sample was obtained as control and placed in a 2-ml vacutainer preconditioned with EDTA on ice. All blood sampling was followed by port and catheter flushing with heparinized saline. The animal was then placed on a heating pad (37 °C) on a surgery table.
  • the animal received a dextrose/saline i.v. infusion (approximately 30 ml/kg/hr) from the catheter that had previously been used for the E-2078 injection.
  • Supplemental ketamine (approximately 5 mg/kg) was administered at hourly intervals to maintain the animal in an anesthetized state throughout the sampling period.
  • E-2078 was dissolved in sale (10 mg/kg in 20 mg/ml solution), and injected in one of the leg catheters (injection time was approximately 15 seconds). At the end of injection, a timer was started, and the injection catheter was flushed with heparinized saline. Blood sampling was performed at the following time points from the contralateral catheter: 0; 5; 15; 30; 60; 90; 120 and 180 minutes. Samples were centrifuged (3,400 x g at 0-5°C for 5 minutes). A 0.2-ml aliquot of plasma was placed in a cryovial containing 1.8 ml of 1 % TFA and kept at -40 °C until the time of analysis.
  • the instrument inco ⁇ orates an HY-400 Nd-YAG laser (Lumonics Inc., Kanata, Ontario, Canada) operated at a wavelength of 355 nm, a pulse duration of ca, 10 ns, and a pulse energy density of ca. 15 mJ/cm 2 ; a 30 KV ion acceleration potential; and a 2 m long linear time-of-flight mass analyzer.
  • Ion detection and signal amplification are accomplished with a hybrid microchannel plate detector- discrete dynode electron multiplier assembly (detector potential being 3.6 KV).
  • the samples with acetonitrile [2: 1 (v/v)] were mixed with matrix solution.
  • a saturated matrix solution was prepared by dissolving excess amount (5 mg/ml of ⁇ -cyano-4-hydroxycinnamic acid in acetonitrile and 0.1 % aqueous TFA [1:2 (v/v)].
  • Plasma containing E-2078 biotransformation products was mixed with matrix solution in a ratio of 1:2 (v/v).
  • a 0.5- ⁇ l aliquot of the resulting solution was applied to the mass spectrometer sample probe tip, and allowed to evaporate to dryness int he air. The sample probe was inserted into the mass spectrometer vacuum system where solvents in the sample were completely removed.
  • a standard curve was constructed using E-2078 standard solutions.
  • the standard solutions were made by serial dilution of E-2078 in 0.05% TFA aqueous solution.
  • Dyn A (1-8) ( ⁇ l of 136 ⁇ g/ml, or 0.136 ⁇ g), was added to 100 ⁇ l of standard solutions.
  • the E-2078 standard solutions containing Dyn A (1-8) were each mixed with acetonitrile (2: 1 , v/v), and the resulting solutions were separately mixed 4HCCA matrix solution (1 :2, v/v).
  • Half- microliter aliquots of the resulting solutions were applied to the mass spectrometer sample probe tip for MALDI MS analysis.
  • Mass spectrometric measurements were carried out in triplicate for each aliquot of the standard solutions. The ratio of the signal intensity from E-2078 to that from the internal standard, and the S.E. of the triplicate measurements were calculated.
  • Dyn A (1-8) on in vitro incubation at 37°C in freshly drawn human and rhesus monkey blood, was rapidly processed to form two biotransformation products: Dyn A (2-8) and Dyn A (1-6).
  • Dyn A (1-8) and the biotransformation products were cleared from human blood at 90 minutes incubation, and cleared from the rhesus monkey blood at 30-minute incubation.
  • E-2078 was also incubated in freshly drawn human and rhesus monkey blood at 37°C for up to 24 hours. In stark contrast to the results observed with Dyn A (1-8), no major biotransformation products were detected. At 24-hour incubation time point, intact E-2078 was still the major species present in the blood.
  • the first linear section from 0 to 10 minutes, represented a rapid distribution of the peptide out of the vascular system into various tissues after injection.
  • the second linear section from 15 to 180 minutes, represented the elimination of E-2078 from both the blood and the tissues, once the distribution had been completed and the plasma and tissue concentration were in equilibrium.
  • the modified Dyn A (1-8), E-2078, did not exhibit significant biotransformation either in vitro in freshly drawn human and rhesus monkey blood, or in vivo in rhesus monkey blood.
  • Modification with a methyl group at Tyr 1 , N-methyl-Tyr 1 effectively protected the cleavage at the N-terminal Tyr 1 position. No biotransformation products from this processing were detected.
  • Peptide linkage of Arg(6)-Arg(7) is usually the other site of cleavage of Dyn A peptides in a biological matrix. Modification with a methyl group at the N-Arg 7 position effectively blocked this biotransformation.
  • Peptide biotransformation may be tissue specific. Reported stability studies on E- 2078 in comparison with Dyn A (1-17) have shown that no E-2078 degradation product was detected after incubation with mouse serum (10%) for 2 hours, whereas Dyn A (1-17) was degraded completely in that time period. However, E- 2078 was biotransformed in mice brain homogenate (1 %) with a half-life about 4 hours; Dyn A (1-17) biotransformation half-life was about 0.5 hours.
  • the cleavage sites of E-2078 in mouse brain homogenates were identified to be Gly(3)- Phe(4), Phe(4)-Leu(5), and Leu(5)-Arg(6) by high-performance liquid chromatography. In this study, no biotransformation products from Gly(3)-Phe(4) cleavage were detected.
  • Dyn A (1-8) analog, E-2078 was very stable in vitro in freshly drawn human and rhesus monkey blood and in vivo in rhesus monkey blood. Minor processing products from E-2078, such as E(l-4), E(l-5) and E(3-6), were detected in vitro in some human and rhesus monkey blood. The apparent half-life of elimination of E-2078 in vivo from the rhesus monkey blood was about 44.0 minutes. Based on these studies, E-2078 is a suitable candidate for development as a therapeutic agent for opioid addiction and cocaine addiction.
  • N-methyl-Tyr',N-methyl-Arg 7 -D-Leu 8 dyno ⁇ hin A (1-8) ethylamide is a dyno ⁇ hin A (1-8) analog.
  • this synthetic peptide is stable against enzymatic cleavages in biological matrices. Modification with a methyl group at Tyr 1 , N-methyl-Tyr 1 , effectively protected the cleavage at the N-terminal Tyr 1 position.
  • Peptide linkage of Arg(6)-Arg(7) is usually the other site of cleavage of dyno ⁇ hin A peptides in a biological matrix.
  • E-2078 binds to kappa opioid receptors just like dyno ⁇ hin A (1-17) as studied in vitro with isolated organ preparations: guinea pig ileum, mouse vas deferens, and rabbit vas deferens. Intravenously administered E-2078 was approximately equipotent to mo ⁇ hine in the tail-pinch assay in mice. This systemic effectiveness of E-2078 was attributed to its stability against enzymatic degradation. In preliminary studies in humans, E-2078 exhibited analgesic properties when used in patients with severe pain after lower abdominal surgery in clinical studies.
  • opioid withdrawal Many signs and symptoms of opioid withdrawal are thought to be mediated primarily by central mechanisms, whereas analgesic or antinociceptive effects of opioids can be either centrally or peripherally mediated, depending on the experimental situation. It is thought that the primary action of cocaine in drug abuse, as well as the molecular abnormalities caused by cocaine in experimental animals are mediated in specific brain regions, thus probably requiring a therapeutic intervention that is centrally targeted.
  • the general concept that some peptides can cross the blood-brain barrier as intact molecules has gained more acceptance in recent years.
  • MALDI MS matrix-assisted laser deso ⁇ tion/ionization mass spectrometry
  • N-methyl-Tyr 1 , N-methyl-Arg 7 -D-Leu 8 Dyn A (1-8) ethylamide (E-2078) was synthesized and kindly supplied by Eisai Co. Ltd. (Ibaraki, Japan).
  • Dyn A (1-8) was purchased from Peninsula Laboratories, Inc. (Belmont, CA).
  • Diphenhydramine HCl was purchased from Sigma Chemical Company (St. Louis, MO).
  • Ketamine HCl was obtained from Fort Dodge (Fort Dodge, IA).
  • Heparin was purchased from Elkins-Sinn (Cherry Hill, NJ). Saline (0.9% NaCl) and dextrose were from Abbott Laboratories (North Chicago, IL).
  • HCCA ⁇ -Cyano-4-hydroxycinnamic acid
  • E-2078 in vivo studies in rhesus monkeys Monkeys were injected with diphenhydramine HCl (1.2 mg/kg, i.m.) as a pretreatment to limit the possible consequences of histamine release following administration of relatively large amounts of E-2078. Thirty minutes later, they were anesthetized with ketamine Hcl (10 mg/kg, i.m.). The back of the lower leg area and the dorsal upper neck/lower skull area were carefully shaved to avoid any adventitious bleeding.
  • An indwelling catheter (Angiocath, 22 gauge, 1 " long, Becton Dickinson, Sandy, UT) was acutely placed in a superficial vein of each leg, secured and flushed with heparinized saline (20 U/ml), and connected to a multi-sample injection port.
  • One mil liter of blood sample was obtained as control and placed in a 2 ml vacutainer preconditioned with EDTA on ice. All blood sampling was followed by port and catheter flushing with heparinized saline. The animal was then placed on a heating pad (7°C) on a surgery table.
  • a spinal needle 22 gauge, 1.5" long.
  • the animal received a dextrose/saline intravenous infusion (approximately 30 ml/kg/hr) from the catheter that had previously been used for the E-2078 injection.
  • Supplemental ketamine HCl (approximately 5 mg/kg) was administered at hourly intervals to maintain the animal in an anesthetized state throughout the sampling period.
  • the required amount of E-2078 was dissolved in saline (10 mg/kg in 20 mg/ml solution).
  • the E-2078 was injected in one of the leg catheters (injection time was approximately 15 seconds).
  • injection time was approximately 15 seconds.
  • a timer was started, and the injection catheter was flushed with heparinized saline. Sampling was performed at the following time points: CSF sampling g, 3, 8, 15, 30, 60, 90, 120, and 180 minutes; blood sampling from the contralateral catheter only at 0, 5, 15, 30, 60, 90, 120, and 180 minutes.
  • the stylet of the spinal needle was removed, and the first CSF drop exiting the spinal needle (approximately 50 ⁇ l) was discarded from each sample.
  • a sample of CSF of 0.1 ml was removed and placed in a 2 ml cryovial containing 0.3 ml of 1 % TFA. The content of the vial was gently mixed and the vial rapidly placed on ice. At all times in the experiments, only CSF samples that did not show signs of blood contamination were used. If blood contamination of the CSF samples was suspected (i.e. , by a slight red coloring in the sample), the spinal needle would be gently moved or removed and re-inserted until a clean sample could be obtained.
  • CSF samples were stored in a -40 °C freezer until the time of analysis. Blood samples were centrifuged (3,400 g at 0-5 °C for 5 minutes). A 0.2 ml aliquot of plasma was placed in a cryovial containing 1.8 ml of 1 % TFA and kept at -40°C until the time of analysis.
  • the plasma samples Prior to analysis, the plasma samples were thawed at room temperature. To 2.0 ml of plasma sample solution was added 40 ⁇ l of 80 ⁇ M (1.54 ⁇ g/ml) Dyn A (1-8) in 0.05% aqueous TFA as the internal standard. The solution containing plasma, the peptide. the internal standard and TFA, was centrifuge-filtered with Centricon-SR3 concentrators (Amicon Inc. , Beverly , MA), with molecular weight cut-off of 3,000 Da. Twenty microliters of the filtrate was mixed with 10 ⁇ l of acetonitrile for analysis by mass spectrometry.
  • Mass spectrometry The samples were analyzed by a mass spectrometer consisting of a matrix-assisted laser deso ⁇ tion ion source, coupled with a linear time-of-flight mass analyzer (MALDI MS). This instrument was constructed at the Rockefeller University (Beavis and Chait, 1989, 1990). A saturated matrix solution was prepared by dissolving excess amount (5 mg/ml) of 4HCCA in acetonitrile and 0.1 % aqueous TFA [1 :2 (v/v)] . For mass spectrometric measurement, the samples with acetonitrile [2: 1 (v/v)] were mixed with the matrix solution.
  • MALDI MS linear time-of-flight mass analyzer
  • CSF or plasma samples were mixed with matrix solution in a ratio of 1 :2 (v/v).
  • a 0.5 ⁇ l aliquot of the resulting solution was applied to the mass spectrometer sample probe tip, and allowed to evaporate to dryness in the air.
  • the sample probe was inserted into the mass spectrometer vacuum system where solvents in the sample were completely removed. After 5 minutes, a working pressure of approximately 10 "7 torr was achieved.
  • the results from 200 laser shots were added to produce each mass spectrum. In this study, one spectrum was obtained for each sample from each subject.
  • a standard curve was constructed using E-2078 standard solutions.
  • the standard solutions were made by serial dilution of E-2078 in 0.05% TFA aqueous solution.
  • a constant amount of internal standard, Dyn A (1-8) (1 ⁇ l of 136 ⁇ g/ml, or 0.136 ⁇ g), was added to 100 ⁇ l of standard solutions.
  • the E-2078 standard solutions containing Dyn A (1-8) were each mixed with acetonitrile (2: 1, v/v), and the resulting solutions were separately mixed with 4HCCA matrix solution (1 :2, v/v).
  • Half microliter aliquots of the resulting solutions were applied to the mass spectrometer sample probe tip for MALDI MS analysis. Mass spectrometric measurements were carried out in triplicate for each aliquot of the standard solutions.
  • Mass spectrometric analysis of CSF from the rhesus monkeys after intravenous injection of E-2078 detected the presence of E-2078 in CSF indicating that E-2078 had crossed the blood-brain barrier .
  • CSF was sampled at the time points, 3, 8, 15, 30, 60, 90, 120 and 180 minutes after E-2078 injection.
  • E-2078 was detected in CSF collected at all the time points from the five rhesus monkey subjects used in the present studies. In the quantitative studies conducted in three monkey subjects, it was found that the signal intensity from E-2078 varied more in CSF than in plasma among the subjects.
  • the concentration of E-2078 in CSF and plasma at various time points was tabulated.
  • the maximum percentage of E-2078 detected in blood relative to the total amount of the peptide injected is about 48%, calculated based upon the reported value of 54 ml/kg blood volume per body weight found in rhesus monkeys.
  • the red blood cell counts (RBC) measured in the CSF from three rhesus monkeys were 0, 0.6 and 1.0 RBC/ ⁇ l, whereas the RBCs of the blood from the same rhesus monkeys were 6.6, 4.7 and 4.0 million RBC/ ⁇ l, respectively.
  • RBC red blood cell counts
  • E-2078 is able to enter CSF following intravenous administration in primates, and is therefore a suitable candidate for the investigation of possible centrally mediated effects of systemically administered dyno ⁇ hins.
  • the Dyn A (1-8) analog, E-2078 was very stable in vivo in rhesus monkey blood. Detection of E-2078 in CSF after intravenous injection indicated that E-2078 had crossed the blood-brain barrier. Based on these studies, E-2078 is a suitable candidate for use as a therapeutic agent for the treatment for specific addictions.

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Abstract

Cette invention concerne un procédé qui permet de réduire les taux de dopamine de base ou d'alterner les poussées des taux de dopamine dans le cerveau d'un mammifère, ce procédé consistant à administrer à un patient nécessitant une telle thérapie une quantité pharmaceutiquement acceptable d'un composé qui est un ligand du récepteur opioïde kappa. Les ligands spécifiques pouvant être utilisés dans cette invention sont la dynorphine A, des fragments actifs ou des analogues de dynorphine A.
PCT/US1997/010916 1996-06-24 1997-06-24 Procede d'utilisation de ligands du recepteur opioïde kappa WO1997049422A1 (fr)

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Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0974363A1 (fr) * 1997-09-02 2000-01-26 Toray Industries, Inc. Remedes contre la toxicomanie
WO2009046868A1 (fr) * 2007-09-11 2009-04-16 Mondobiotech Laboratories Ag Utilisation d'un peptide en tant qu'agent thérapeutique
WO2009046857A1 (fr) * 2007-09-11 2009-04-16 Mondobiotech Laboratories Ag Utilisation d'un peptide en tant qu'agent thérapeutique
WO2020247599A1 (fr) * 2019-06-06 2020-12-10 The Rockefeller University Agonistes pyrano [3,4,b] pyrazine kappa 4-substitués pour le traitement de la pharmacodépendance
US11091497B2 (en) 2017-12-08 2021-08-17 The Rockefeller University Pyrano[3,4-b]pyrazine kappa opioid receptor ligands for treating addiction, pruritus, pain, and inflammation

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WO1995003804A1 (fr) * 1993-07-27 1995-02-09 Albert Einstein College Of Medicine Of Yeshiva University, A Division Of Yeshiva University Procedes d'amplification de l'efficacite analgesique d'opiaces ou de desintoxication d'un individu dependant d'opiaces
WO1996002251A1 (fr) * 1994-07-19 1996-02-01 Albert Einstein College Of Medicine Of Yeshiva University, A Division Of Yeshiva University Procede permettant d'accroitre la puissance analgesique tout en reduisant le potentiel de dependance d'agonistes opioides exogenes et endogenes

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EP0614913A2 (fr) * 1984-11-09 1994-09-14 EISAI Co., Ltd. Dérivés de la dynorphine
WO1995003804A1 (fr) * 1993-07-27 1995-02-09 Albert Einstein College Of Medicine Of Yeshiva University, A Division Of Yeshiva University Procedes d'amplification de l'efficacite analgesique d'opiaces ou de desintoxication d'un individu dependant d'opiaces
WO1996002251A1 (fr) * 1994-07-19 1996-02-01 Albert Einstein College Of Medicine Of Yeshiva University, A Division Of Yeshiva University Procede permettant d'accroitre la puissance analgesique tout en reduisant le potentiel de dependance d'agonistes opioides exogenes et endogenes

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SHIPPENBERG T S ET AL: "Sensitization to the behavioral effects of cocaine: Modulation by dynorphin and kappa- opioid receptor agonists.", PHARMACOLOGY BIOCHEMISTRY AND BEHAVIOR 57 (3). 1997. 449-455, XP002043591 *
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Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0974363A1 (fr) * 1997-09-02 2000-01-26 Toray Industries, Inc. Remedes contre la toxicomanie
EP0974363A4 (fr) * 1997-09-02 2003-05-07 Toray Industries Remedes contre la toxicomanie
WO2009046868A1 (fr) * 2007-09-11 2009-04-16 Mondobiotech Laboratories Ag Utilisation d'un peptide en tant qu'agent thérapeutique
WO2009046857A1 (fr) * 2007-09-11 2009-04-16 Mondobiotech Laboratories Ag Utilisation d'un peptide en tant qu'agent thérapeutique
US11091497B2 (en) 2017-12-08 2021-08-17 The Rockefeller University Pyrano[3,4-b]pyrazine kappa opioid receptor ligands for treating addiction, pruritus, pain, and inflammation
WO2020247599A1 (fr) * 2019-06-06 2020-12-10 The Rockefeller University Agonistes pyrano [3,4,b] pyrazine kappa 4-substitués pour le traitement de la pharmacodépendance

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