Identification and characterization of multiple splice variants of the kappa3-related opioid receptor (KOR-3) gene
OwnerGavril Pasternak, Ying-Xian Paninventors
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Abstract
The present invention encompasses novel splice variant forms of the muopioid receptor-1 (KOR-3) and the polynuclotide sepuences encoding the KOR-3 splice variants. The invention further encompasses methods of screening for compositions regulating the KOR-3 splice variant activities and the development of therapeutic modatities directed to regulating activity. Regulation of the KOR-3 splice varient activities may impact the physiologic processes of analgesia and weight management.
Claims80
An isolated KOR-3 splice variant polypeptide comprising at least five consecutive amino acid residues thereof and retaining KOR-3 activity, where the KOR-3 polypeptides are not the same as any known polypeptides derived from a KOR-3 polypeptide.
An isolated polynucleotide, or a complementary strand thereto that hybridizes under stringent conditions, comprising at least 15 consecutive nucleotides of the KOR-3 polynucleotide fragment depicted in FIGS. 2-8 where the polynucleotide is not the same as a polynucleotide derived from any known KOR-3 polynucleotide.
The polynucleotide according to claim 0.21, where the nucleotide fragment comprises at least 15 consecutive nucleotides of rat KOR-3A as depicted in FIG. 13.
A polynucleotide, or a complementary strand thereto that hybridizes under stringent conditions, of at least 15 consecutive nucleotides encoding the isolated KOR-3 polypeptide.
A method of screening compositions for opioid activity comprising the steps of
- a) obtaining a control cell that does not express a recombinant opioid receptor;
- b) obtaining a test cell that is the same as the control cell except that it expresses a recombinant opioid receptor selected from the group consisting of an KOR-3 splice variant polypeptide;
- c) contacting the control cell and test cell with an amount of an opioid sufficient to exert a physiologic effect;
- d) separately measuring the physiologic effect of the composition on the control cell and test cell;
- and
- e) comparing the physiologic effect of the composition to the physiologic effect of the opioid, where determination of a physiologic effect of the composition is expressed relative to that of the opioid.
A method of screening compositions for opioid activity comprising the steps of
- a) obtaining a control polypeptide that is not a recombinant opioid receptor;
- b) obtaining a test polypeptide that is a recombinant opioid receptor selected from the group consisting of an KOR-3 splice variant polypeptide;
- c) contacting a composition with the control polypeptide and the test polypeptide;
- d) contacting the test polypeptide with an amount of an opioid sufficient to measurably bind the test polypeptide;
- c) measuring the binding of the composition and the opioid;
- and
- e) comparing test polypeptide binding of the composition to that of the opioid, where determination of binding of the composition is expressed relative to that of the opioid.
A method of screening compositions for differential opioid activity comprising the steps of
- a) obtaining a first test polypeptide selected from the group consisting of an KOR-3 splice variant polypeptide, and contacting it with a composition;
- b) obtaining a second test polypeptide selected from the group consisting of an KOR-3 splice variant polypeptide;
- c) measuring the binding of the composition to the first and second test polypeptides;
- and
- d) comparing the binding of the composition and the first test polypeptide to that of the second test polypeptide where differential activity is expressed as a ratio of the two binding affinities.
A non-human animal in which one or both endogenous KOR-3 alleles has been altered by homologous recombination with an exogenously introduced nucleic acid.
A non-human transgenic animal carrying a transgene comprising a nucleic acid that encodes a polypeptide selected from the group consisting of an KOR-3 splice variant polypeptide.
A method for regulating morphine analgesia in a subject comprising altering the amount of KOR-3 polypeptide activity by
- a) administering antigen binding fragments to a subject in an amount of and a duration sufficient to regulate morphine analgesia;
- or
- b) administering agonists to a subject in an amount of and a duration sufficient to regulate morphine analgesia;
- or
- c) administering antagonists to a subject in an amount of and a duration sufficient to regulate morphine analgesia;
- or
- d) administering small molecule ligands to a subject in an amount of and a duration sufficient to regulate morphine analgesia;
- or
- e) administering a DNA plasmid vector containing a nucleic acid encoding a polypeptide selected from the group consisting of an KOR-3 splice variant polypeptide, thereby expressing an KOR-3 splice variant polypeptide in a subject in an amount of and a duration sufficient to regulate morphine analgesia;
- or
- f) administering an antisense nucleic acid corresponding to a nucleic acid comprising a polypeptide encoding a polypeptide selected from the group consisting of an KOR-3 splice variant polypeptide, to a subject in an amount of and a duration sufficient to regulate morphine analgesia.
- and wherein the antigen binding fragment, agonist, antagonist or small molecule ligand is directed to a polypeptide selected from the group consisting of an KOR-3 splice variant polypeptide.
A method for regulating body weight in a subject comprising altering the level of KOR-3 polypeptide activity by
- a) administering antigen binding fragments to a subject in an amount of and a duration sufficient to regulate body weight;
- or
- b) administering agonists to a subject in an amount of and duration sufficient to regulate body weight;
- or
- c) administering antagonists to a subject in an amount of and a duration sufficient to regulate body weight;
- or
- d) administering small molecule ligands to a subject in an amount of and a duration sufficient to regulate body weight;
- or
- e) administering a DNA plasmid vector containing a nucleic acid encoding a polypeptide selected from the group consisting of an KOR-3 splice variant polypeptide, thereby expressing an KOR-3 splice variant polypeptide in a subject in an amount of and a duration sufficient to regulate body weight;
- or
- f) administering an antisense nucleic acid corresponding to a nucleic acid encoding a polypeptide selected from the group consisting of an KOR-3 splice variant polypeptide, to a subject in an amount of and a duration sufficient to regulate body weight.
- and wherein the antigen binding fragment, agonist, antagonist or small molecule ligand is directed to a polypeptide selected from the group consisting of an KOR-3 splice variant polypeptide.
A method for diagnosing an KOR-3 splice variant-associated pharmacological abnormality, comprising measuring the amount of variant activity or tissue distribution thereof in a subject and comparing that activity or tissue distribution to a control sample, wherein a difference in the amount of activity or tissue distribution correlates with the presence of a pharmacological defect.
A method for diagnosing an KOR-3 splice variant-associated disorder of the neuroendocrine system, comprising measuring the amount of variant activity or tissue distribution in a subject and comparing that activity or tissue distribution to a control sample, wherein a difference in the amount of activity or tissue distribution correlates with the presence of a defect within the neuroendocrine system.
A method for generating antigen binding fragments specific for a KOR-3 splice variant polypeptide.
Dependencies are inferred from the claim text.