The Salk Institute for Biological Studies

Research Development Foundation

University of California

University of Utah Research Foundation

Other

Scripps Clinic and Research Foundation

Jean E Rivier

Wylie W Vale, Jr

Catherine L Rivier

John S Porter

Joachim Spiess

Marvin R Brown

Carl A Hoeger

Marilyn H Perrin

Kathy A Lewis

Wayne D Kornreich

Baldomero M Olivera

Joan Vaughan

Paula G Theobald

Lourdes J Cruz

Jozsef Gulyas

Koichi Kunitake

Jean-Francois Hernandez

Wolfgang H Fischer

John Dykert

Harry A Anderson

Francoise G Presse

Jean-Louis M Nahon

Ronald Mark Evans

David R Hillyard

Michael G Rosenfeld

Neal A Scott

John B Hogenesch

William R Gray

Paul E Sawchenko

Joan M Vaughan

Teresa M Reyes

Chris E Hopkins

Josep L Torres

Fe Abogadie

Karl P Lederis

Jaroslav H Boublik

Guangcheng Jiang

Botond Penke

Steven C Koerber

Arnold T Hagler

Nicholas C Ling

Maren Watkins

Wylie Walker Vale, Jr

R Tyler McCabe

Roger C Guillemin

Frederick S Esch

Richard T Layer

Li-Ming Zhou

Doju Yoshikami

Roger W Roeske

Craig S Walker

Ki-Joon Shon

Gregory S Shen

Elsie C Jimenez

Shao-Yao Ying

Judit Erchegyi

Helmut R Maecke

Roland Riek

Antonio De Miranda

Clark Colledge

Nancy M Sherwood

Wolfgang Fischer

Keith L MacCannell

Dean A Kirby

Cindy Donaldson

Jeffrey Schwartz

Joan Vaughn

Jean F Hernandez

Fe C Abogadie

Million Mulugeta

Robert F Galyean

Walter E Marki

Michael J Brownstein

Susan G Amara

Lee E Eiden

Larry Swanson

Janos I Varga

R Scott Struthers

Kelly E Mayo

Gail Cerelli

Lixin Wang

Anne Corrigan

Richard A McClintock

Grace Christy Rani Royappa

Yvette Taché

Denis McMaster

Alice L Yu

John Yu

C Wayne Bardin

Koichi S Kunitake

Jean Claude Reubi

Graeme Semple

This disclosure reveals that cyclic corticotrophin releasing factor (CRF) antagonist peptides (such as astressin B, its functional fragments, and their derivatives) induce hair growth and prevent hair loss in vivo. This important discovery enables, for instance, methods of promoting hair growth, and methods of treating hair loss (such as the hair loss that occurs normally in some individuals or that is the result of a health disorder or therapeutic treatment). Exemplary cyclic CRF antagonist peptides useful in the disclosed methods are provided throughout the disclosure.

Methods for promoting hair growth

A search of the public human genome database identified a human EST, GenBank accession number AW293249, which has high homology to known pufferfish urocortin sequences. The full length sequence was amplified from human genomic DNA and sequenced. Sequence homology comparisons of the novel sequence with human urocortin I and urocortin II revealed that the sequence encoded a novel human urocortin, which was designated urocortin III (UcnIII). While urocortin III does not have high affinity for either CRF-R1 or CRF-R2, the affinity for CRF-R2 is greater than the affinity for CRF-R1. Urocortin III is capable stimulating cyclic AMP production in cells expressing CRF-R2&#x3b1; or &#x3b2;. Thus, the affinity is high enough that urocortin III could act as a native agonist of CRF-R2. However, it is also likely that urocortin III is a stronger agonist of a yet to be identified receptor.

Urocortin-III and uses thereof

A human urocortin-related peptide with significant sequence homology to the CRF neuropeptide family was identified. A mouse cDNA was isolated from whole brain poly (A+) RNA that encodes a predicted 38 amino acid peptide protein designated herein as urocortin II. Both human URP and mouse Ucn II are structurally related to the other known mammalian family members, CRF and urocortin (Ucn). These peptides are involved in the regulation of the hypothalamic-pituitary-adrenal axis under basal and stress conditions, suggesting a similar role for URP and Ucn IL Synthesized Ucn-II and URP peptide binds with higher affinity to CRF-R2 than to CRF-R1 Ucn II and human URP appear to be involved in the regulation of body temperature and appetite and may play a role in other stress related phenomenon. These findings identify Ucn II and human URP as a new members of the CRF family of neuropeptides, which are expressed centrally and bind to CRF-R2.

Urocortin proteins and uses thereof

The present invention relates to a method for identifying modulators of B1 G-protein coupled receptors. The present invention also relates to a method for identifying an antagonist or agonist of the corticotropin-releasing factor receptor 2 (CRFR2). The present invention also relates to a method for improving antagonists or agonists of CRFR2. The present invention also relates to the three-dimensional structure of CRFR2 as representative of the B1 GPCR subfamily and its use as a basis for rational drug design of antagonist or agonists of B1 GPCRs.

Structure-based modulators of B1 G-protein coupled receptors

A human urocortin-related peptide with significant sequence homology to the CRF neuropeptide family was identified. A mouse cDNA was isolated from whole brain poly (A+) RNA that encodes a predicted 38 amino acid peptide protein designated herein as urocortin II. Both human URP and mouse Ucn II are structurally related to the other known mammalian family members, CRF and urocortin (Ucn). These peptides are involved in the regulation of the hypothalamic-pituituary-adrenal axis under basal and stress conditions, suggesting a similar role for URP and Ucn II. Synthesized Ucn-II and URP peptide binds with higher affinity to CRF-R2 than to CRF-R1 Ucn II and human URP appear to be involved in the regulation of body temperature and appetite and may play a role in other stress related phenomenon. These findings identify Ucn II and human URP as a new members of the CRF family of neuropeptides, which are expressed centrally and bind to CRF-R2.

The present invention is directed to conantokin peptides, conantokin peptide derivatives and conantokin peptide chimeras, referred to collectively as conantokins, having 10-30 amino acids, including preferably two or more &ggr;-carboxyglutamic acid residues. The conantokins are useful for the treatment of neurologic and psychiatric disorders, such as anticonvulsant agents, neuroprotective agents or analgesic agents.

Conantokins

Substantially pure conotoxin peptides are provided which inhibit synaptic transmissions at the neuromuscular junctions and which are useful both in vivo and in assays because they specifically target particular skeletal nAChRs to the exclusion of neuronal nAChRs. The peptides are of such length that they can be made by chemical synthesis, and the preferred peptides have formula: H-His-4Hyp-4Hyp-Cys-Cys-Leu-Tyr-Gly-Lys-Cys-Arg-Arg-Tyr-4Hyp-Gly-Cys-Ser-S er-Ala-Ser-Cys-Cys-Gln-Xaa.sub.24 -NH.sub.2 wherein Xaa.sub.24 is Arg or Gly.

Conotoxin peptides

Novel cyclic CRF antagonist peptides have the amino acid sequence: ##STR1## wherein Y is Ac, H, Ac-Thr or H-Thr; R.sub.30 is Glu or Cys; R.sub.32 is His or preferably a basic and/or aromatic D-amino acid such as D-His or D-Arg; R.sub.33 is Lys, Orn or Cys. The N-terminus may be extended by Leu or Asp-Leu. CML may be substituted for Leu.sup.27, and D-Tyr may be substituted for D-Phe to facilitate labelling. Lys may be substituted for Arg.sup.23, and its side chain connected by a lactam bridge to Glu.sup.20 to form a bicyclic peptide. Disclosed CRF antagonists include:

Cyclic CRF antagonists

Novel cyclic CRF agonist peptides have the amino acid sequence: (cyclo 30-33)Ac-Pro-Pro-Ile-Ser-Leu-Asp-Leu-Thr-D-Phe-His-Leu-Leu-Arg-Glu-Val-Leu -Glu-Nle-Ala-Arg-Ala-Glu-Gln-Leu-Ala-Gln-R.sub.30 -Ala-R.sub.32 -R.sub.33 -Asn-Arg-Lys-Leu-Nle-Glu-Ile-Ile-NH.sub.2 wherein R.sub.30 is Glu or Cys; R.sub.32 is His or a D-amino acid such as D-His, D-Arg or similar; R.sub.33 is Lys, Orn or Cys. The N-terminus may be extended by Ser-Glu-Glu. Lys may be substituted for Arg.sup.23, and its side chain connected by a lactam bridge to Glu.sup.20 to form a bicyclic peptide. Certain disclosed CRF agonists include:

Cyclic CRF agonists

Novel cyclic CRF agonist peptides have the amino acid sequence: (cyclo 30-33)Ac-Pro-Pro-Ile-Ser-Leu-Asp-Leu-Thr-D-Phe-His-Leu-Leu-Arg-Glu-Val-Leu -Glu-Nle-Ala-Arg-Ala-Glu-Gln-Leu-Ala-Gln-R.sub.30 -Ala-R.sub.32 -R.sub.33 -Asn-Arg-Lys-Leu-Nle-Glu-Ile-Ile-NH.sub.2 wherein R.sub.30 is Glu or Cys; R.sub.32 is His, D-His or an equivalent .alpha.-amino acid; R.sub.33 is Lys, Orn or Cys. The N-terminus may be extended by Ser-Glu-Glu or shortened by up to three more residues. Lys may be substituted for Arg.sup.23, and its side chain connected by a lactam bridge to Glu.sup.20 to form a bicyclic peptide. Certain disclosed CRF agonists include:

Betides having at least one betidamino acid are provided which have GnRH antagonistic properties. These antagonists may be used to regulate fertility and to treat steroid-dependent tumors, such as prostatic and mammary tumors. One particularly effective betide, which is an analog of the decapeptide GnRH that is highly soluble in water at physiologic pH, has the formula: Ac-D-2Nal-D-4Cpa-D-3Pal-Ser-Aph(acetyl)-D-Aph(acetyl)-Leu-D/L-Agl(methyl,i sopropyl .beta.-alanyl)-Pro-D-Ala-NH.sub.2, and another has the formula Ac-D-2Nal-D-Cpa-D-3Pal-D/L-Agl(For)-Aph(Ac)-D-Aph(Ac)-Leu-ILys-Pro-D-Ala-N H.sub.2.

GNRH antagonist betides

Compounds termed 'betides' mimic peptides and contain one or more residues of aminoglycine, C.sup..alpha. -aminoalanine, aminosarcosine or the like wherein the side chain amino group has been acylated and optionally also alkylated. Generally, betides have the formula: X.sub.N -X.sub.1 -X.sub.2 -X.sub.3 -X.sub.m -X.sub.4 -X.sub.5 -X.sub.6 -X.sub.C, where X.sub.N is an acyl or other N-terminal group or a peptide up to about 50 amino acids in length having such a group; X.sub.C is OH, NH.sub.2 or other C-terminal group or a peptide up to about 50 amino acids in length having such a group; and X.sub.1 -X.sub.6 are each independently a betidamino acid or .alpha.-amino acid or des-X; and X.sub.m is a peptide up to about 50 amino acids or des-X; provided however that at least one of X.sub.1 -X.sub.6 is a betidamino acid residue having the formula: ##STR1## wherein R.sub.0 is H or CH.sub.3, R and R.sub.2 are H or lower alkyl, and R.sub.3 is an acyl group, an isocyanate or isothiocyanate group, a sulfonyl group or the like. To make a betide, an aminoglycine residue is subjected to side chain acylation, and optionally also alkylation, after it is coupled into a peptide intermediate. By synthesizing betides with multiple substituents at one or more positions in an otherwise peptidic chain, efficient screening of betides which mimic peptides having a large number of different natural or unnatural amino acid substituents at a particular position, and optionally both D- and L-isomers thereof, is possible.

Methods of making and screening betide libraries

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Jean E Rivier